Hairy-root organ cultures for the production of human acetylcholinesterase

PubMed Central

Woods, Ryan R; Geyer, Brian C; Mor, Tsafrir S

2008-01-01

Background Human cholinesterases can be used as a bioscavenger of organophosphate toxins used as pesticides and chemical warfare nerve agents. The practicality of this approach depends on the availability of the human enzymes, but because of inherent supply and regulatory constraints, a suitable production system is yet to be identified. Results As a promising alternative, we report the creation of "hairy root" organ cultures derived via Agrobacterium rhizogenes-mediated transformation from human acetylcholinesterase-expressing transgenic Nicotiana benthamiana plants. Acetylcholinesterase-expressing hairy root cultures had a slower growth rate, reached to the stationary phase faster and grew to lower maximal densities as compared to wild type control cultures. Acetylcholinesterase accumulated to levels of up to 3.3% of total soluble protein, ~3 fold higher than the expression level observed in the parental plant. The enzyme was purified to electrophoretic homogeneity. Enzymatic properties were nearly identical to those of the transgenic plant-derived enzyme as well as to those of mammalian cell culture derived enzyme. Pharmacokinetic properties of the hairy-root culture derived enzyme demonstrated a biphasic clearing profile. We demonstrate that master banking of plant material is possible by storage at 4°C for up to 5 months. Conclusion Our results support the feasibility of using plant organ cultures as a successful alternative to traditional transgenic plant and mammalian cell culture technologies. PMID:19105816

Targeting Mutant BRAF with Vemurafenib in Relapsed or Refractory Hairy Cell Leukemia

PubMed Central

Tiacci, Enrico; Park, Jae H.; De Carolis, Luca; Chung, Stephen S.; Broccoli, Alessandro; Scott, Sasinya; Zaja, Francesco; Devlin, Sean; Pulsoni, Alessandro; Chung, Young Rock; Cimminiello, Michele; Kim, Eunhee; Rossi, Davide; Stone, Richard M.; Motta, Giovanna; Saven, Alan; Varettoni, Marzia; Altman, Jessica K.; Anastasia, Antonella; Grever, Michael R.; Ambrosetti, Achille; Rai, Kanti R.; Fraticelli, Vincenzo; Lacouture, Mario E.; Carella, Angelo Michele; Levine, Ross L.; Leoni, Pietro; Rambaldi, Alessandro; Falzetti, Franca; Ascani, Stefano; Capponi, Monia; Martelli, Maria Paola; Park, Christopher Y.; Pileri, Stefano Aldo; Rosen, Neal; Foà, Robin; Berger, Michael F.; Zinzani, Pier Luigi; Abdel-Wahab, Omar; Falini, Brunangelo; Tallman, Martin S.

2016-01-01

BACKGROUND BRAF-V600E is the genetic lesion underlying hairy cell leukemia. We assessed the safety and activity of the oral BRAF inhibitor vemurafenib in patients with hairy cell leukemia who relapsed after or were refractory to purine analogues. METHODS We conducted in Italy and USA two phase-2 single-arm multicenter studies of vemurafenib (960 mg twice daily) given for a median of 16 and 18 weeks, respectively. Primary endpoints were complete remission rate and overall response rate. Patient enrollment was completed (n=28) in the Italian trial in April 2013 and is still open (n=26/36) in the American trial. RESULTS Drug-related adverse events were usually of grade 1-2, and those most frequently requiring dose reductions were rash and arthralgia/arthritis; secondary cutaneous tumors (treated with simple excision) developed in 6/50 patients. Overall response rates were 96% (25/26 evaluable Italian patients) and 100% (24/24 evaluable American patients), obtained after a median of 8 weeks and 12 weeks, respectively. Complete response rates were 34.6% (9/26) and 41.7% (10/24), respectively. In the Italian trial, after a median follow-up of 23 months, the median relapse-free and treatment-free survivals were respectively 19 and 25 months in complete responders, and 6 and 18 months in partial responders. In the American trial, 1-year progression-free and overall survival were 73% and 91%, respectively. Frequent persistence of phospho-ERK+ bone marrow leukemic cells at the end of treatment suggests bypass MEK-ERK reactivation as a resistance mechanism. CONCLUSIONS A short oral course of vemurafenib proved safe and highly effective in relapsed/refractory hairy cell leukemia patients (Funded by AIRC, ERC, Roche/Genentech and others; EudractCT number: 2011-005487-13, ClinicalTrials.gov number NCT01711632). PMID:26352686

Leukemic meningitis in a patient with hairy cell leukemia. A case report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wolfe, D.W.; Scopelliti, J.A.; Boselli, B.D.

1984-09-15

Central nervous system involvement has not previously been described in patients with hairy cell leukemia (HCL). A patient is reported who presented with meningeal involvement as his initial symptom of HCL. Diagnosis was established by morphologic and cytochemical studies of his cerebrospinal fluid (CSF) and bone marrow. Treatment with whole-brain irradiation and intrathecal chemotherapy was successful in clearing leukemic cells from the CSF with resolution of symptoms.

Targeting Mutant BRAF in Relapsed or Refractory Hairy-Cell Leukemia.

PubMed

Tiacci, Enrico; Park, Jae H; De Carolis, Luca; Chung, Stephen S; Broccoli, Alessandro; Scott, Sasinya; Zaja, Francesco; Devlin, Sean; Pulsoni, Alessandro; Chung, Young R; Cimminiello, Michele; Kim, Eunhee; Rossi, Davide; Stone, Richard M; Motta, Giovanna; Saven, Alan; Varettoni, Marzia; Altman, Jessica K; Anastasia, Antonella; Grever, Michael R; Ambrosetti, Achille; Rai, Kanti R; Fraticelli, Vincenzo; Lacouture, Mario E; Carella, Angelo M; Levine, Ross L; Leoni, Pietro; Rambaldi, Alessandro; Falzetti, Franca; Ascani, Stefano; Capponi, Monia; Martelli, Maria P; Park, Christopher Y; Pileri, Stefano A; Rosen, Neal; Foà, Robin; Berger, Michael F; Zinzani, Pier L; Abdel-Wahab, Omar; Falini, Brunangelo; Tallman, Martin S

2015-10-29

BRAF V600E is the genetic lesion underlying hairy-cell leukemia. We assessed the safety and activity of the oral BRAF inhibitor vemurafenib in patients with hairy-cell leukemia that had relapsed after treatment with a purine analogue or who had disease that was refractory to purine analogues. We conducted two phase 2, single-group, multicenter studies of vemurafenib (at a dose of 960 mg twice daily)--one in Italy and one in the United States. The therapy was administered for a median of 16 weeks in the Italian study and 18 weeks in the U.S. study. Primary end points were the complete response rate (in the Italian trial) and the overall response rate (in the U.S. trial). Enrollment was completed (28 patients) in the Italian trial in April 2013 and is still open (26 of 36 planned patients) in the U.S. trial. The overall response rates were 96% (25 of 26 patients who could be evaluated) after a median of 8 weeks in the Italian study and 100% (24 of 24) after a median of 12 weeks in the U.S. study. The rates of complete response were 35% (9 of 26 patients) and 42% (10 of 24) in the two trials, respectively. In the Italian trial, after a median follow-up of 23 months, the median relapse-free survival was 19 months among patients with a complete response and 6 months among those with a partial response; the median treatment-free survival was 25 months and 18 months, respectively. In the U.S. trial, at 1 year, the progression-free survival rate was 73% and the overall survival rate was 91%. Drug-related adverse events were usually of grade 1 or 2, and the events most frequently leading to dose reductions were rash and arthralgia or arthritis. Secondary cutaneous tumors (treated with simple excision) developed in 7 of 50 patients. The frequent persistence of phosphorylated ERK-positive leukemic cells in bone marrow at the end of treatment suggests bypass reactivation of MEK and ERK as a resistance mechanism. A short oral course of vemurafenib was highly effective in

Real-time detection of BRAF V600E mutation from archival hairy cell leukemia FFPE tissue by nanopore sequencing.

PubMed

Vacca, Davide; Cancila, Valeria; Gulino, Alessandro; Lo Bosco, Giosuè; Belmonte, Beatrice; Di Napoli, Arianna; Florena, Ada Maria; Tripodo, Claudio; Arancio, Walter

2018-02-01

The MinION is a miniaturized high-throughput next generation sequencing platform of novel conception. The use of nucleic acids derived from formalin-fixed paraffin-embedded samples is highly desirable, but their adoption for molecular assays is hurdled by the high degree of fragmentation and by the chemical-induced mutations stemming from the fixation protocols. In order to investigate the suitability of MinION sequencing on formalin-fixed paraffin-embedded samples, the presence and frequency of BRAF c.1799T > A mutation was investigated in two archival tissue specimens of Hairy cell leukemia and Hairy cell leukemia Variant. Despite the poor quality of the starting DNA, BRAF mutation was successfully detected in the Hairy cell leukemia sample with around 50% of the reads obtained within 2 h of the sequencing start. Notably, the mutational burden of the Hairy cell leukemia sample as derived from nanopore sequencing proved to be comparable to a sensitive method for the detection of point mutations, namely the Digital PCR, using a validated assay. Nanopore sequencing can be adopted for targeted sequencing of genetic lesions on critical DNA samples such as those extracted from archival routine formalin-fixed paraffin-embedded samples. This result let speculating about the possibility that the nanopore sequencing could be trustably adopted for the real-time targeted sequencing of genetic lesions. Our report opens the window for the adoption of nanopore sequencing in molecular pathology for research and diagnostics.

Xanthones from roots, hairy roots and cell suspension cultures of selected Hypericum species and their antifungal activity against Candida albicans.

PubMed

Zubrická, Daniela; Mišianiková, Anna; Henzelyová, Jana; Valletta, Alessio; De Angelis, Giulia; D'Auria, Felicia Diodata; Simonetti, Giovanna; Pasqua, Gabriella; Čellárová, Eva

2015-11-01

Highest xanthone contents were found in Hypericum pulchrum and H. annulatum untransformed roots. The best anti- Candida activity was obtained for hairy roots extracts of H. tetrapterum clone 2 ATCC 15834. Extracts of root cultures, hairy roots and cell suspensions of selected Hypericum spp. were screened for the presence of xanthones and tested for their antifungal activity against Candida albicans strain ATCC 10231. At least one of the following xanthones, 5-methoxy-2-deprenylrheediaxanthone; 1,3,6,7-tetrahydroxyxanthone; 1,3,5,6-tetrahydroxyxanthone; paxanthone; kielcorin or mangiferin was identified in methanolic extracts of the untransformed root cultures. The highest total xanthone content, with five xanthones, was found in untransformed H. pulchrum and H. annulatum root cultures. Hairy roots and the controls of H. tetrapterum contained 1,7-dihydroxyxanthone, while hairy root cultures and the corresponding controls of H. tomentosum contained toxyloxanthone B, 1,3,6,7- and 1,3,5,6-tetrahydroxyxanthone. Two xanthones, cadensin G and paxanthone, were identified in cell suspension cultures of H. perforatum. Their content increased about two-fold following elicitation with salicylic acid. The anti-Candida activity of the obtained extracts ranged from MIC 64 to >256 µg ml(-1). Among the extracts of Hypericum untransformed roots, the best antifungal activity was obtained for extracts of H. annulatum grown under CD conditions. Extracts of hairy roots clones A4 and 7 ATCC15834 of H. tomentosum and clone 2 ATCC15834 of H. tetrapterum displayed inhibition of 90% of Candida growth with 256 μg ml(-1). Extracts from chitosan-elicitated cells did not show antifungal activity.

Accumulation of cell wall-bound phenolic metabolites and their upliftment in hairy root cultures of tomato (Lycopersicon esculentum Mill.).

PubMed

Mandal, Sudhamoy; Mitra, Adinpunya

2008-07-01

Alkaline hydrolysis of cell wall material of tomato hairy roots yielded ferulic acid as the major phenolic compound. Other phenolics were 4-hydroxybenzoic acid, vanillic acid, 4-hydroxybenzaldehyde, vanillin and 4-coumaric acid. The content of phenolics was much higher at the early stage of hairy root growth. The ferulic acid content decreased up to 30 days and then sharply increased to 360 microg/g at 60 days of growth. Elicitation of hairy root cultures with Fusarium mat extract (FME) increased ferulic acid content 4-fold after 24 h. As the pathogen-derived elicitors have specific receptors in plants, FME may thus be used for inducing resistance against Fusarium oxysporum f. sp. lycopersici.

Metabolism of oxybenzone in a hairy root culture: Perspectives for phytoremediation of a widely used sunscreen agent.

PubMed

Chen, Feiran; Huber, Christian; May, Robert; Schröder, Peter

2016-04-05

Oxybenzone (OBZ), known as Benzophenone-3, is a commonly used UV filter in sun tans and skin protectants, entering aquatic systems either directly during recreational activities or indirectly through wastewater treatment plants discharge. To study the potential degradation capacity of plants for OBZ in phytotreatment, a well-established hairy root culture (Armoracia rusticana) was treated with OBZ. More than 20% of spiked OBZ (100μM) was eliminated from the medium by hairy roots after 3h of exposure. Two metabolites were identified as oxybenzone-glucoside (OBZ-Glu) and oxybenzone-(6-O-malonyl)-glucoside (OBZ-Mal-Glu) by LC-MS/MS and TOF-MS. Formation of these metabolites was confirmed by enzymatic synthesis, as well as enzymatic and alkaline hydrolysis. Incubation with O-glucosyltransferase (O-GT) extracted from roots formed OBZ-Glu; whereas β-d-Glucosidase hydrolyzed OBZ-Glu. However, alkaline hydrolysis led to cleavage of OBZ-Mal-Glu and yielded OBZ-Glu. In the hairy root culture, an excretion of OBZ-Glu into the growth medium was observed while the corresponding OBZ-Mal-Glu remained stored in root cells over the incubation time. We propose that metabolism of oxybenzone in plants involves initial conjugation with glucose to form OBZ-Glu followed by malonylation to yield OBZ-Mal-Glu. To our best knowledge this first finding presenting the potential of plants to degrade benzophenone type UV filters by phytoremediation. Copyright © 2015 Elsevier B.V. All rights reserved.

Exploring the Metabolic Stability of Engineered Hairy Roots after 16 Years Maintenance.

PubMed

Häkkinen, Suvi T; Moyano, Elisabeth; Cusidó, Rosa M; Oksman-Caldentey, Kirsi-Marja

2016-01-01

Plants remain a major source of new drugs, leads and fine chemicals. Cell cultures deriving from plants offer a fascinating tool to study plant metabolic pathways and offer large scale production systems for valuable compounds - commercial examples include compounds such as paclitaxel. The major constraint with undifferentiated cell cultures is that they are generally considered to be genetically unstable and cultured cells tend to produce low yields of secondary metabolites especially over time. Hairy roots, a tumor tissue caused by infection of Agrobacterium rhizogenes is a relevant alternative for plant secondary metabolite production for being fast growing, able to grow without phytohormones, and displaying higher stability than undifferentiated cells. Although genetic and metabolic stability has often been connected to transgenic hairy roots, there are only few reports on how a very long-term subculturing effects on the production capacity of hairy roots. In this study, hairy roots producing high tropane alkaloid levels were subjected to 16-year follow-up in relation to genetic and metabolic stability. Cryopreservation method for hairy roots of Hyoscyamus muticus was developed to replace laborious subculturing, and although the post-thaw recovery rates remained low, the expression of transgene remained unaltered in cryopreserved roots. It was shown that although displaying some fluctuation in the metabolite yields, even an exceedingly long-term subculturing was successfully applied without significant loss of metabolic activity.

Novel therapeutic options for relapsed hairy cell leukemia.

PubMed

Jain, Preetesh; Polliack, Aaron; Ravandi, Farhad

2015-01-01

The majority of patients with hairy cell leukemia (HCL) achieve a response to therapy with cladribine or pentostatin with or without rituximab. However, late relapses can occur. Treatment of relapsed HCL can be difficult due to a poor tolerance to chemotherapy, increased risk of infections and decreased responsiveness to chemotherapy. The identification of BRAFV600E mutations and the role of aberrant MEK kinase and Bruton's tyrosine kinase (BTK) pathways in the pathogenesis of HCL have helped to develop novel targeted therapies for these patients. Currently, the most promising therapeutic strategies for relapsed or refractory HCL include recombinant immunoconjugates targeting CD22 (e.g. moxetumomab pasudotox), BRAF inhibitors such as vemurafenib and B cell receptor signaling kinase inhibitors such as ibrutinib. Furthermore, the VH4-34 molecular variant of classic HCL has been identified to be less responsive to chemotherapy. Herein, we review the results of the ongoing clinical trials and potential future therapies for relapsed/refractory HCL.

BRAF inhibition in hairy cell leukemia with low-dose vemurafenib.

PubMed

Dietrich, Sascha; Pircher, Andreas; Endris, Volker; Peyrade, Frédéric; Wendtner, Clemens-Martin; Follows, George A; Hüllein, Jennifer; Jethwa, Alexander; Ellert, Elena; Walther, Tatjana; Liu, Xiyang; Dyer, Martin J S; Elter, Thomas; Brummer, Tilman; Zeiser, Robert; Hermann, Michael; Herold, Michael; Weichert, Wilko; Dearden, Claire; Haferlach, Torsten; Seiffert, Martina; Hallek, Michael; von Kalle, Christof; Ho, Anthony D; Gaehler, Anita; Andrulis, Mindaugas; Steurer, Michael; Zenz, Thorsten

2016-06-09

The activating mutation of the BRAF serine/threonine protein kinase (BRAF V600E) is the key driver mutation in hairy cell leukemia (HCL), suggesting opportunities for therapeutic targeting. We analyzed the course of 21 HCL patients treated with vemurafenib outside of trials with individual dosing regimens (240-1920 mg/d; median treatment duration, 90 days). Vemurafenib treatment improved blood counts in all patients, with platelets, neutrophils, and hemoglobin recovering within 28, 43, and 55 days (median), respectively. Complete remission was achieved in 40% (6/15 of evaluable patients) and median event-free survival was 17 months. Response rate and kinetics of response were independent of vemurafenib dosing. Retreatment with vemurafenib led to similar response patterns (n = 6). Pharmacodynamic analysis of BRAF V600E downstream targets showed that vemurafenib (480 mg/d) completely abrogated extracellular signal-regulated kinase phosphorylation of hairy cells in vivo. Typical side effects also occurred at low dosing regimens. We observed the development of acute myeloid lymphoma (AML) subtype M6 in 1 patient, and the course suggested disease acceleration triggered by vemurafenib. The phosphatidylinositol 3-kinase hotspot mutation (E545K) was identified in the AML clone, providing a potential novel mechanism for paradoxical BRAF activation. These data provide proof of dependence of HCL on active BRAF signaling. We provide evidence that antitumor and side effects are observed with 480 mg vemurafenib, suggesting that dosing regimens in BRAF-driven cancers could warrant reassessment in trials with implications for cost of cancer care. © 2016 by The American Society of Hematology.

[Induction of hairy roots of Panax ginseng and studies on suitable culture condition of ginseng hairy roots].

PubMed

Zhao, Shou-Jing; Li, Chang-Yu; Qian, Yan-Chun; Luo, Xiao-Pei; Zhang, Xin; Wang, Xue-Song; Kang, Bo-Yu

2004-03-01

Ginseng is a valuable medicinal plant with ginsenosides as its mian effective components. Because ginseng is a perennial plant and has a very strict demand for soil conditions, the way of cultivating ginseng by cutting woods is still used in China at present and thus forest resources has been extremely destroyed. Increasing attention has been paid to the hairy roots induced by the infection of Agrobacterium rhizogenes in the production of plant secondary metabolic products for the hairy roots are characterized by rapid growth and stable hereditary and biochemical traits. That has opened a new way for the industrial production of ginseosides. However, there is little report for such studies from China. In this paper, hairy roots of ginseng were induced from the root explants of two-year-old ginseng by Agrobacterium rhizogenes A4 with directly inoculating. The transformed hairy roots could grow rapidly on MS medium and 1/2 MS medium without hormones. The cultured clones of the hairy roots were established on a solid 1/2 MS medium. After 4 - 5 subcultures the hairy roots still maintained a vigorous growth. A pair of primers were designed and synthesized according to the analytical results of RiA4TL-DNA sequence by Slightom et al . 0.8kb rolC was obtained by PCR using the genome DNA of hairy root of ginseng. Transformation was confirmed by PCR amplification of rolC genes from the hairy roots of P. ginseng. Growth rate of hairy roots on liquid medium increased by 2 times then that of the solid medium. The growth of the hairy roots can be divided into three stages: high speed in the first two weeks, middle speed in the 3 - 4 weeks and low speed hereafter. Changing the culture solution at 2 weeks regular intervals is conductive to maintaining the rapid growth of the hairy roots. By means of determination for specific growth rate and ginsenosides content, the high-yield hairy root clone R9923 was selected. The content of monomer gisenoside of Rg1, Re, Rf, Rbl, Rc, Rb2 and

Differentials on graph complexes II: hairy graphs

NASA Astrophysics Data System (ADS)

Khoroshkin, Anton; Willwacher, Thomas; Živković, Marko

2017-10-01

We study the cohomology of the hairy graph complexes which compute the rational homotopy of embedding spaces, generalizing the Vassiliev invariants of knot theory. We provide spectral sequences converging to zero whose first pages contain the hairy graph cohomology. Our results yield a way to construct many nonzero hairy graph cohomology classes out of (known) non-hairy classes by studying the cancellations in those sequences. This provide a first glimpse at the tentative global structure of the hairy graph cohomology.

Implant-retained skull prosthesis to cover a large defect of the hairy skull resulting from treatment of a basal cell carcinoma: A clinical report.

PubMed

Hoekstra, Jitske; Vissink, Arjan; Raghoebar, Gerry M; Visser, Anita

2017-05-01

Skin carcinoma, particularly basal cell carcinoma, and its treatment can result in large defects of the hairy skull. A 53-year-old man is described who was surgically treated for a large basal cell carcinoma invading the skin and underlying tissue at the top of the hairy skull. Treatment consisted of resecting the tumor and external part of the skull bone. To protect the brain and to cover the defect of the hairy skull, an acrylic resin skull prosthesis with hair was designed to mask the defect. The skull prosthesis was retained on 8 extraoral implants placed at the margins of the defect in the skull bone. The patient was satisfied with the treatment outcome. Copyright © 2016 Editorial Council for the Journal of Prosthetic Dentistry. Published by Elsevier Inc. All rights reserved.

Persistent Legionnaire's disease in an adult with hairy cell leukemia successfully treated with prolonged levofloxacin therapy.

PubMed

Cunha, Burke A; Munoz-Gomez, Sigridh; Gran, Arthur; Raza, Muhammad; Irshad, Nadia

2015-01-01

Legionnaire's disease (LD) manifests most commonly as an atypical community acquired pneumonia (CAP) with systemic extrapulmonary manifestations. Disorders associated with impaired cell mediated immunity (CMI) are particularly predisposed to LD. Hairy cell leukemia (HCL) is a rare B-cell lymphoproliferative leukemia associated with decreased CMI. LD has only rarely been reported in HCL. We present a most interesting case of persistent LD in a elderly male with HCL who required prolonged antibiotic therapy. Copyright © 2015 Elsevier Inc. All rights reserved.

Rituximab with pentostatin or cladribine: an effective combination treatment for hairy cell leukemia after disease recurrence.

PubMed

Else, Monica; Dearden, Claire E; Matutes, Estella; Forconi, Francesco; Lauria, Francesco; Ahmad, Humayun; Kelly, Susan; Liyanage, Anandika; Ratnayake, Vijitha; Shankari, Jagadeesan; Whalley, Ioana; Catovsky, Daniel

2011-06-01

The purine analogs pentostatin and cladribine are effective treatments for hairy cell leukemia (HCL). However, alternative treatments are needed for patients with recurrent disease. We reviewed retrospectively data from 18 patients who were retreated with either pentostatin (n = 12) or cladribine (n = 6) in combination with rituximab, after 1-6 (median 2) previous treatments with either purine analog as a single agent. All 18 patients responded to therapy, with a complete response (CR) rate of 89%. This compared favorably with CR rates of 68% after second-line therapy and 47% after third-line therapy in 88 patients retreated one or more times with a purine analog alone. Toxicity with the combination treatment was minimal. At a median follow-up of 36 months (range 5-83 months) all 16 complete responders remained in CR, while one partial responder developed recurrent disease at 10 months. The estimated recurrence rate at 3 years was 7%. This compares with 21% after second-line therapy and 42% after third-line therapy in the 88 patients retreated with a purine analog alone. Furthermore, it was a marked improvement on the 55% recurrence at 3 years previously seen in these same 18 patients after their own first-line treatment with single-agent pentostatin or cladribine (p = 0.006). The combination of a purine analog with rituximab was safe and effective for patients with recurrent HCL. The results suggest an added benefit compared with single-agent purine analog therapy.

Biologically active recombinant human erythropoietin expressed in hairy root cultures and regenerated plantlets of Nicotiana tabacum L.

PubMed Central

Schäfer, Holger; Ramamoorthy, Siva; Wink, Michael

2017-01-01

Hairy root culture is a potential alternative to conventional mammalian cell culture to produce recombinant proteins due to its ease in protein recovery, low costs and absence of potentially human pathogenic contaminants. The current study focussed to develop a new platform of a hairy root culture system from Nicotiana tabacum for the production of recombinant human EPO (rhEPO), which is regularly produced in mammalian cells. The human EPO construct was amplified with C-terminal hexahistidine tag from a cDNA of Caco-2 cells. Two versions of rhEPO clones, with or without the N-terminal calreticulin (cal) fusion sequence, were produced by cloning the amplified construct into gateway binary vector pK7WG2D. Following Agrobacterium rhizogenes mediated transformation of tobacco explants; integration and expression of constructs in hairy roots were confirmed by several tests at DNA, RNA and protein levels. The amount of intracellular rhEPO from hairy root cultures with cal signal peptide was measured up to 66.75 ng g-1 of total soluble protein. The presence of the ER signal peptide (cal) was essential for the secretion of rhEPO into the spent medium; no protein was detected from hairy root cultures without ER signal peptide. The addition of polyvinylpyrrolidone enhanced the stabilization of secreted rhEPO leading to a 5.6 fold increase to a maximum concentration of 185.48 pg rhEPOHR g-1 FW hairy root cultures. The rhizo-secreted rhEPO was separated by HPLC and its biological activity was confirmed by testing distinct parameters for proliferation and survival in retinal pigment epithelial cells (ARPE). In addition, the rhEPO was detected to an amount 14.8 ng g-1 of total soluble leaf protein in transgenic T0 generation plantlets regenerated from hairy root cultures with cal signal peptide. PMID:28800637

Transgenic hairy roots. recent trends and applications.

PubMed

Giri, A; Narasu, M L

2000-03-01

Agrobacterium rhizogenes causes hairy root disease in plants. The neoplastic roots produced by A. rhizogenes infection is characterized by high growth rate and genetic stability. These genetically transformed root cultures can produce higher levels of secondary metabolites or amounts comparable to that of intact plants. Hairy root cultures offer promise for production of valuable secondary metabolites in many plants. The main constraint for commercial exploitation of hairy root cultures is their scaling up, as there is a need for developing a specially designed bioreactor that permits the growth of interconnected tissues unevenly distributed throughout the vessel. Rheological characteristics of heterogeneous system should also be taken into consideration during mass scale culturing of hairy roots. Development of bioreactor models for hairy root cultures is still a recent phenomenon. It is also necessary to develop computer-aided models for different parameters such as oxygen consumption and excretion of product to the medium. Further, transformed roots are able to regenerate genetically stable plants as transgenics or clones. This property of rapid growth and high plantlet regeneration frequency allows clonal propagation of elite plants. In addition, the altered phenotype of hairy root regenerants (hairy root syndrome) is useful in plant breeding programs with plants of ornamental interest. In vitro transformation and regeneration from hairy roots facilitates application of biotechnology to tree species. The ability to manipulate trees at a cellular and molecular level shows great potential for clonal propagation and genetic improvement. Transgenic root system offers tremendous potential for introducing additional genes along with the Ri T-DNA genes for alteration of metabolic pathways and production of useful metabolites or compounds of interest. This article discusses various applications and perspectives of hairy root cultures and the recent progress achieved

Azadirachtin production by hairy root cultivation of Azadirachta indica in a modified stirred tank reactor.

PubMed

Srivastava, Smita; Srivastava, A K

2012-11-01

Present investigation involves hairy root cultivation of Azadirachta indica in a modified stirred tank reactor under optimized culture conditions for maximum volumetric productivity of azadirachtin. The selected hairy root line (Az-35) was induced via Agrobacterium rhizogenes LBA 920-mediated transformation of A. indica leaf explants (Coimbatore variety, India). Liquid culture of the hairy roots was developed in a modified Murashige and Skoog medium (MM2). To further enhance the productivity of azadirachtin, selected growth regulators (1.0 mg/l IAA and 0.025 mg/l GA(3)), permeabilizing agent (0.5 % v/v DNBP), a biotic elicitor (1 % v/v Curvularia (culture filtrate)) and an indirectly linked biosynthetic precursor (50 mg/l cholesterol) were added in the growth medium on 15th day of the hairy root cultivation period in shake flask. Highest azadirachtin production (113 mg/l) was obtained on 25th day of the growth cycle with a biomass of 21 g/l DW. Further, batch cultivation of hairy roots was carried out in a novel liquid-phase bioreactor configuration (modified stirred tank reactor with polyurethane foam as root support) to investigate the possible scale-up of the established A. indica hairy root culture. A biomass production of 15.2 g/l with azadirachtin accumulation in the hairy roots of 6.4 mg/g (97.28 mg/l) could be achieved after 25 days of the batch cultivation period, which was ~27 and ~14 % less biomass and azadirachtin concentration obtained respectively, in shake flasks. An overall volumetric productivity of 3.89 mg/(l day) of azadirachtin was obtained in the bioreactor.

Black hairy tongue associated with olanzapine treatment: a case report.

PubMed

Tamam, Lut; Annagur, Bilge Burcak

2006-10-01

Olanzapine is an atypical antipsychotic drug approved for acute and long-term treatment of bipolar disorder. Although relatively safe as compared to other classical antipsychotic medications, there are a number of uncommon adverse effects of olanzapine such as oral cavity lesions. In addition to the relatively common side effect of dry mouth, several articles have reported an association between olanzapine treatment and the development of oral lesions such as apthous stomatitis, pharyngitis, glossitis and oral ulceration. Although there are several cases in which the tongue was affected in conjunction with stomatitis or pharyngitis, we could not find a case report indicating a direct relationship between olanzapine use and a tongue lesion. We present here the case of a patient with bipolar disorder, who developed recurrent black hairy tongue on two different occasions following the addition of olanzapine to lithium treatment. In the present case, xerostomia (dry mouth), which is an adverse reaction of both olanzapine and lithium, may have played a role in the development of black hairy tongue. All agents with a possible side effect of xerostomia may predispose patients to black hairy tongue, especially when they are administered in combination. To preclude the development of this complication with such drugs, extra time and effort should be given to improving oral hygiene.

An immunophenotypic and molecular diagnosis of composite hairy cell leukaemia and chronic lymphocytic leukaemia.

PubMed

Liptrot, Stuart; O' Brien, David; Langabeer, Stephen E; Quinn, Fiona; Mackarel, A Jill; Elder, Patrick; Vandenberghe, Elisabeth; Hayden, Patrick J

2013-12-01

Hairy cell leukaemia (HCL) and chronic lymphocytic leukaemia (CLL) are distinct clinicopathological B cell chronic lymphoproliferative disorders (B-CLPD). Both diseases have characteristic immunophenotypic and molecular features. The co-existence of two B-CLPD is perhaps more common than previously thought but a composite HCL and CLL has been rarely documented. A case is reported in which the morphology, integrated with an extensive immunophenotyping panel, and incorporation of the recently described HCL-associated BRAF V600E mutation, enabled the prompt diagnosis of composite HCL and CLL thus allowing appropriate treatment selection. This case serves to highlight the benefit of a multidisciplinary approach to the diagnosis of bi-clonal B-CLPD.

Scalarized hairy black holes

NASA Astrophysics Data System (ADS)

Kleihaus, Burkhard; Kunz, Jutta; Yazadjiev, Stoytcho

2015-05-01

In the presence of a complex scalar field scalar-tensor theory allows for scalarized rotating hairy black holes. We exhibit the domain of existence for these scalarized black holes, which is bounded by scalarized rotating boson stars and hairy black holes of General Relativity. We discuss the global properties of these solutions. Like their counterparts in general relativity, their angular momentum may exceed the Kerr bound, and their ergosurfaces may consist of a sphere and a ring, i.e., form an ergo-Saturn.

Cost-benefit analysis of interferon alfa-2b in treatment of hairy cell leukemia.

PubMed

Ozer, H; Golomb, H M; Zimmerman, H; Spiegel, R J

1989-04-19

The clinical benefits as well as the cost benefits of use of recombinant interferon (IFN) alfa-2b instead of conventional chemotherapy (primarily chlorambucil) for progressive hairy cell leukemia were assessed retrospectively on the basis of 12 months of clinical data from 128 patients treated with IFN alfa-2b. Data from 71 matched historical control patients who had received conventional treatment were used for survival analysis. Hematologic response (reversal of cytopenias) was achieved by 18% of the control patients versus 73% of the IFN-treated patients. This response was associated with virtual elimination of the need for transfusions and splenectomy as well as dramatic decreases in the frequency of fatal infections (22.5% vs. 1.6%) and the 12-month mortality rate (28% vs. 3.1%). Direct costs per patient per year for medical care (transfusions, antibiotic treatment, splenectomy, and chemotherapy) of those receiving IFN alfa-2b were 2.8-fold lower than costs for medical care of control patients ($5,027 vs. $14,046). Indirect costs, which reflect the present value of future earnings lost due to premature death, were 13.3-fold lower for IFN-treated patients than for control patients ($4,771 vs. $63,507). Our analysis demonstrates that IFN alfa-2b offers substantial clinical and cost advantages to patients with hairy cell leukemia and that the introduction of this therapy using novel biotechnology furthers the health care community's commitment to cost containment.

Consensus guidelines for the diagnosis and management of patients with classic hairy cell leukemia.

PubMed

Grever, Michael R; Abdel-Wahab, Omar; Andritsos, Leslie A; Banerji, Versha; Barrientos, Jacqueline; Blachly, James S; Call, Timothy G; Catovsky, Daniel; Dearden, Claire; Demeter, Judit; Else, Monica; Forconi, Francesco; Gozzetti, Alessandro; Ho, Anthony D; Johnston, James B; Jones, Jeffrey; Juliusson, Gunnar; Kraut, Eric; Kreitman, Robert J; Larratt, Loree; Lauria, Francesco; Lozanski, Gerard; Montserrat, Emili; Parikh, Sameer A; Park, Jae H; Polliack, Aaron; Quest, Graeme R; Rai, Kanti R; Ravandi, Farhad; Robak, Tadeusz; Saven, Alan; Seymour, John F; Tadmor, Tamar; Tallman, Martin S; Tam, Constantine; Tiacci, Enrico; Troussard, Xavier; Zent, Clive S; Zenz, Thorsten; Zinzani, Pier Luigi; Falini, Brunangelo

2017-02-02

Hairy cell leukemia is an uncommon hematologic malignancy characterized by pancytopenia and marked susceptibility to infection. Tremendous progress in the management of patients with this disease has resulted in high response rates and improved survival, yet relapse and an appropriate approach to re-treatment present continuing areas for research. The disease and its effective treatment are associated with immunosuppression. Because more patients are being treated with alternative programs, comparison of results will require general agreement on definitions of response, relapse, and methods of determining minimal residual disease. The development of internationally accepted, reproducible criteria is of paramount importance in evaluating and comparing clinical trials to provide optimal care. Despite the success achieved in managing these patients, continued participation in available clinical trials in the first-line and particularly in the relapse setting is highly recommended. The Hairy Cell Leukemia Foundation convened an international conference to provide common definitions and structure to guide current management. There is substantial opportunity for continued research in this disease. In addition to the importance of optimizing the prevention and management of the serious risk of infection, organized evaluations of minimal residual disease and treatment at relapse offer ample opportunities for clinical research. Finally, a scholarly evaluation of quality of life in the increasing number of survivors of this now manageable chronic illness merits further study. The development of consensus guidelines for this disease offers a framework for continued enhancement of the outcome for patients.

Consensus guidelines for the diagnosis and management of patients with classic hairy cell leukemia

PubMed Central

Abdel-Wahab, Omar; Andritsos, Leslie A.; Banerji, Versha; Barrientos, Jacqueline; Blachly, James S.; Call, Timothy G.; Catovsky, Daniel; Dearden, Claire; Demeter, Judit; Else, Monica; Forconi, Francesco; Gozzetti, Alessandro; Ho, Anthony D.; Johnston, James B.; Jones, Jeffrey; Juliusson, Gunnar; Kraut, Eric; Kreitman, Robert J.; Larratt, Loree; Lauria, Francesco; Lozanski, Gerard; Montserrat, Emili; Parikh, Sameer A.; Park, Jae H.; Polliack, Aaron; Quest, Graeme R.; Rai, Kanti R.; Ravandi, Farhad; Robak, Tadeusz; Saven, Alan; Seymour, John F.; Tadmor, Tamar; Tallman, Martin S.; Tam, Constantine; Tiacci, Enrico; Troussard, Xavier; Zent, Clive S.; Zenz, Thorsten; Zinzani, Pier Luigi; Falini, Brunangelo

2017-01-01

Hairy cell leukemia is an uncommon hematologic malignancy characterized by pancytopenia and marked susceptibility to infection. Tremendous progress in the management of patients with this disease has resulted in high response rates and improved survival, yet relapse and an appropriate approach to re-treatment present continuing areas for research. The disease and its effective treatment are associated with immunosuppression. Because more patients are being treated with alternative programs, comparison of results will require general agreement on definitions of response, relapse, and methods of determining minimal residual disease. The development of internationally accepted, reproducible criteria is of paramount importance in evaluating and comparing clinical trials to provide optimal care. Despite the success achieved in managing these patients, continued participation in available clinical trials in the first-line and particularly in the relapse setting is highly recommended. The Hairy Cell Leukemia Foundation convened an international conference to provide common definitions and structure to guide current management. There is substantial opportunity for continued research in this disease. In addition to the importance of optimizing the prevention and management of the serious risk of infection, organized evaluations of minimal residual disease and treatment at relapse offer ample opportunities for clinical research. Finally, a scholarly evaluation of quality of life in the increasing number of survivors of this now manageable chronic illness merits further study. The development of consensus guidelines for this disease offers a framework for continued enhancement of the outcome for patients. PMID:27903528

Air entrainment in hairy surfaces

NASA Astrophysics Data System (ADS)

Nasto, Alice; Regli, Marianne; Brun, P.-T.; Alvarado, José; Clanet, Christophe; Hosoi, A. E.

2016-07-01

Motivated by diving semiaquatic mammals, we investigate the mechanism of dynamic air entrainment in hairy surfaces submerged in liquid. Hairy surfaces are cast out of polydimethylsiloxane elastomer and plunged into a fluid bath at different velocities. Experimentally, we find that the amount of air entrained is greater than what is expected for smooth surfaces. Theoretically, we show that the hairy surface can be considered as a porous medium and we describe the air entrainment via a competition between the hydrostatic forcing and the viscous resistance in the pores. A phase diagram that includes data from our experiments and biological data from diving semiaquatic mammals is included to place the model system in a biological context and predict the regime for which the animal is protected by a plastron of air.

Surface topography of hairy cell leukemia cells compared to other leukemias as seen by scanning electron microscopy.

PubMed

Polliack, Aaron; Tadmor, Tamar

2011-06-01

This short review deals with the ultrastructural surface architecture of hairy cell leukemia (HCL) compared to other leukemic cells, as seen by scanning electron microscopy (SEM). The development of improved techniques for preparing blood cells for SEM in the 1970s readily enabled these features to be visualized more accurately. This review returns us to the earlier history of SEM, when the surface topography of normal and neoplastic cells was visualized and reported for the first time, in an era before the emergence and use of monoclonal antibodies and flow cytometry, now used routinely to define cells by their immunophenotype. Surface microvilli are characteristic for normal and leukemic lymphoid cells, myelo-monocytic cells lack microvilli and show surface ruffles, while leukemic plasma and myeloma cells and megakaryocytes display large surface blebs. HCL cell surfaces are complex and typically 'hybrid' in nature, displaying both lymphoid and monocytic features with florid ruffles of varying sizes interspersed with clumps of short microvilli cytoplasm. The surface features of other leukemic cells and photomicrographs of immuno-SEM labeling of cells employing antibodies and colloidal gold, reported more than 20 years ago, are shown.

Drinking with a hairy tongue: viscous entrainment by dipping hairy surfaces

NASA Astrophysics Data System (ADS)

Nasto, Alice; Brun, Pierre-Thomas; Alvarado, José; Bush, John; Hosoi, Anette

2016-11-01

Nectar-drinking bats have tongues covered with hair-like papillae, enhancing their ability to take up viscous nectar by dipping. Using a combination of model experiments and theory reminiscent of Landau-Levich-Derjaguin dip coating, we rationalize this mechanism of viscous entrainment in a hairy texture. For the model experiments, hairy surfaces are fabricated using laser cut molds and casting samples with PDMS elastomer. Modeling the liquid trapped within the texture using a Darcy-Brinkman like approach, we derive the drainage flow solution. The amount of fluid that is entrained is dependent on the viscosity of the fluid, the density of the hairs, and the dipping speed. We find that there is an optimal hair density to maximize fluid uptake.

Three-dimensional measurement of yarn hairiness via multiperspective images

NASA Astrophysics Data System (ADS)

Wang, Lei; Xu, Bugao; Gao, Weidong

2018-02-01

Yarn hairiness is one of the essential parameters for assessing yarn quality. Most of the currently used yarn measurement systems are based on two-dimensional (2-D) photoelectric measurements, which are likely to underestimate levels of yarn hairiness because hairy fibers on a yarn surface are often projected or occluded in these 2-D systems. A three-dimensional (3-D) test method for hairiness measurement using a multiperspective imaging system is presented. The system was developed to reconstruct a 3-D yarn model for tracing the actual length of hairy fibers on a yarn surface. Five views of a yarn from different perspectives were created by two angled mirrors and simultaneously captured in one panoramic picture by a camera. A 3-D model was built by extracting the yarn silhouettes in the five views and transferring the silhouettes into a common coordinate system. From the 3-D model, curved hair fibers were traced spatially so that projection and occlusion occurring in the current systems could be avoided. In the experiment, the proposed method was compared with two commercial instruments, i.e., the Uster Tester and Zweigle Tester. It is demonstrated that the length distribution of hairy fibers measured from the 3-D model showed an exponential growth when the fiber length is sorted from shortest to longest. The hairiness measurements, such as H-value, measured by the multiperspective method were highly consistent with those of Uster Tester (r=0.992) but had larger values than those obtained from Uster Tester and Zweigle Tester, proving that the proposed method corrected underestimated hairiness measurements in the commercial systems.

Oil-soluble hairy nanoparticles as lubricant additives

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhao, Bin

Oil-soluble polymer brush-grafted nanoparticles (hairy NPs) were synthesized by surface-initiated atom transfer radical polymerization of lauryl methacrylate from initiator-functionalized silica nanoparticles and used as an additive for polyalphaolefin (PAO) for friction and wear reduction. Addition of 1 wt% hairy nanoparticles into PAO led to significant friction and wear reduction compared with PAO base oil.

Hairy cell leukaemia complicated by anti-MAG paraproteinemic demyelinating neuropathy: resolution of neurological syndrome after cladribrine treatment.

PubMed

Rossi, Davide; Franceschetti, Silvia; Cerri, Michaela; Conconi, Annarita; Lunghi, Monia; Capello, Daniela; Cantello, Roberto; Gaidano, Gianluca

2007-06-01

Hairy cell leukaemia (HCL) occasionally displays a monoclonal gammopathy, yet the association of HCL with paraproteinemic demyelinating neuropathy (PDN) has not been reported. We describe a HCL case complicated by PDN and high titers of monoclonal IgM against myelin associated glycoprotein (MAG). Heavy and light chains of the patient's anti-MAG monoclonal protein were consistent with those expressed by HCL cells. After treatment with cladribrine, remission of HCL strictly paralleled disappearance of the IgM monoclonal protein and of the serum anti-MAG activity, and led to PDN clinical and electrophysiological improvement. Purine analogs may represent a choice in IgM PDN associated with lymphoproliferative disorders.

Production of glucosinolates, phenolic compounds and associated gene expression profiles of hairy root cultures in turnip (Brassica rapa ssp. rapa).

PubMed

Chung, Ill-Min; Rekha, Kaliyaperumal; Rajakumar, Govindasamy; Thiruvengadam, Muthu

2016-12-01

Turnip (Brassica rapa ssp. rapa) is an important vegetable crop producing glucosinolates (GSLs) and phenolic compounds. The GSLs, phenolic compound contents and transcript levels in hairy root cultures, as well as their antioxidant, antimicrobial and anticancer activity were studied in turnip. Transgenic hairy root lines were confirmed by polymerase chain reaction (PCR) and reverse transcription-PCR. GSLs levels (glucoallysin, glucobrassicanapin, gluconasturtiin, glucobrassicin, 4-methoxyglucobrassicin, neoglucobrassicin and 4-hydroxyglucobrassicin) and their gene expression levels (BrMYB28, BrMYB29, BrMYB34, BrMYB51, BrMYB122, CYP79 and CYP83) significantly increased in hairy roots compared with that in non-transformed roots. Furthermore, hairy roots efficiently produced several important individual phenolic compounds (flavonols, hydroxybenzoic and hydroxycinnamic acids). Colorimetric analysis revealed that the highest levels of total phenol, flavonoid contents, and their gene expression levels (PAL, CHI and FLS) in hairy roots than non-transformed roots. Our study provides beneficial information on the molecular and physiological active processes that are associated with the phytochemical content and biosynthetic gene expression in turnip. Moreover, antioxidant activity, as measured by DPPH scavenging activity, reducing potential, phosphomolybdenum and ferrous ion chelating ability assays was significantly higher in hairy roots. Hairy root extracts exhibited higher antimicrobial activity against bacterial and fungal species. The extract of hairy roots showed inhibition of human breast and colon cancer cell lines.

Hernandulcin in hairy root cultures of Lippia dulcis.

PubMed

Sauerwein, M; Yamazaki, T; Shimomura, K

1991-02-01

The hairy root culture of Lippia dulcis Trev., Verbenaceae, was established by transformation with Agrobacterium rhizogenes A4. The transformed roots grew well in Murashige and Skoog medium containing 2% sucrose. The roots turned light green when they were cultured under 16 h/day light. The green hairy roots produced the sweet sesquiterpene hernandulcin (ca. 0.25 mg/g dry wt) together with 20 other mono- and sesquiterpenes, while no terpenes were detected in the nontransformed root cultures. The growth and hernandulcin production in the hairy root cultures were influenced by the addition of auxins to the medium. The addition of a low concentration of chitosan (0.2 - 10.0 mg / l) enhanced the production of hernandulcin 5-fold.

Drag reduction of a hairy disk

NASA Astrophysics Data System (ADS)

Niu, Jun; Hu, David L.

2011-10-01

We investigate experimentally the hydrodynamics of a hairy disk immersed in a two-dimensional flowing soap film. Drag force is measured as a function of hair length, density, and coating area. An optimum combination of these parameters yields a drag reduction of 17%, which confirms previous numerical predictions (15%). Flow visualization indicates the primary mechanism for drag reduction is the bending, adhesion, and reinforcement of hairs trailing the disk, which reduces wake width and traps "dead water." Thus, the use of hairy coatings can substantially reduce an object's drag while negligibly increasing its weight.

Light requirement for shoot regeneration in horseradish hairy roots.

PubMed

Saitou, T; Kamada, H; Harada, H

1992-08-01

Hairy roots of horseradish (Armoracia rusticana) were induced by inoculation with Agrobacterium rhizogenes harboring Ri plasmid and cultured on phytohormone-free Murashige and Skoog medium after eliminating the bacteria. Hairy roots grew vigorously and sometimes formed yellowish calli under dark conditions. On the other hand, growth of hairy roots stopped after several weeks of culture with light, then shoots were regenerated. Frequency of shoot formation from hairy roots increased as the culture period in light lengthened and the light intensity increased. The shoot regeneration was induced by treatment with white or red light, but not with far-red light. Shoot regeneration by red light was inhibited by following treatment with far-red light. Red and far-red light reversibly affected shoot regeneration. Excised roots of nontransformed plants grew quite slowly on phytohormone-free Murashige and Skoog medium and occasionally formed shoots under white light conditions.

Rotating hairy black holes.

PubMed

Kleihaus, B; Kunz, J

2001-04-23

We construct stationary black-hole solutions in SU(2) Einstein-Yang-Mills theory which carry angular momentum and electric charge. Possessing nontrivial non-Abelian magnetic fields outside their regular event horizon, they represent nonperturbative rotating hairy black holes.

Viscous entrainment on hairy surfaces

NASA Astrophysics Data System (ADS)

Nasto, Alice; Brun, P.-T.; Hosoi, A. E.

2018-02-01

Nectar-drinking bats and honeybees have tongues covered with hairlike structures, enhancing their ability to take up viscous nectar by dipping. Using a combination of model experiments and theory, we explore the physical mechanisms that govern viscous entrainment in a hairy texture. Hairy surfaces are fabricated using laser cut molds and casting samples with polydimethylsiloxane (PDMS) elastomer. We model the liquid trapped within the texture using a Darcy-Brinkmann-like approach and derive the drainage flow solution. The amount of fluid that is entrained is dependent on the viscosity of the fluid, the density of the hairs, and the withdrawal speed. Both experiments and theory reveal an optimal hair density to maximize fluid uptake.

Hairy root culture optimization and resveratrol production from Vitis vinifera subsp. sylvesteris.

PubMed

Hosseini, Sayed Mehdi; Bahramnejad, Bahman; Douleti Baneh, Hamed; Emamifar, Aryo; Goodwin, Paul H

2017-04-01

Resveratrol is a polyphenolic compound produced in very low levels in grapes. To achieve high yield of resveratrol in wild grape, three Agrobacterium rhizogenes strains, Ar318, ArA4 and LBA9402, were used to induce hairy roots following infection of internodes, nodes or petioles of in vitro grown Vitis vinifera subsp. sylvesteris accessions W2 and W16, and cultivar Rasha. The effects of inoculation time, age of explants, bacterial concentration and co-cultivation times were examined on the efficiency of the production of hairy roots. Strains Ar318, ArA4 and LBA9402 all induced hairy roots in the tested genotypes, but the efficiency of ArA4 strain was higher than the other strains. The highest hairy root production was with using internodes as explants. The transformation of hairy roots lines was confirmed by PCR detection of rolB gene. Half Murashige and Skoog (MS) medium was better for biomass production compared with MS medium. HPLC analysis of resveratrol production in the hairy root cultures showed that all the genotypes produced higher amounts of resveratrol than control roots. The highest amount of resveratrol was produced from W16 internode cultures, which was 31-fold higher than that of control root. Furthermore, TLC analysis showed that treatments of hairy roots with sodium acetate and jasmonate elevated resveratrol levels both in hairy root tissue and excreted into the half MS medium. These results demonstrate that endogenous and exogenous factors can affect resveratrol production in hairy root culture of grape, and this strategy could be used to increase low resveratrol production in grapes.

Thymol derivatives from hairy roots of Arnica montana.

PubMed

Weremczuk-Jezyna, I; Kisiel, W; Wysokińska, H

2006-09-01

Five known thymol derivatives were isolated from roots of Arnica montana transformed with Agrobacterium rhizogenes LBA 9402. The compounds were characterized by spectral methods. The pattern of thymol derivatives in light-grown hairy roots was slightly different from that in dark-grown ones. This is the first report on the presence of thymol derivatives in hairy roots of the plant.

Combating photooxidative stress in green hairy roots of Daucus carota cultivated under light irradiation.

PubMed

Mukherjee, Chiranjit; Sircar, Debabrata; Chatterjee, Moniya; Das, Sampa; Mitra, Adinpunya

2014-01-15

The light-dependent generation of active oxygen species, which can disrupt normal metabolic process of plant, is termed as photo-oxidative stress. Plants are equipped with enzymatic and non-enzymatic antioxidative defence system to reduce the effect of such stress. Hairy root culture of Daucus carota when cultivated under continuous illumination (250 μmol m(-2)s(-1)) turned green. To know the reason behind that and photo-oxidative stress response in green hairy roots, activities of several antioxidant enzymes were measured. When compared with normal hairy roots, green hairy roots showed an enhanced superoxide dismutase (SOD) activity. Treatment with a SOD inhibitor diethyldithiocarbamate led to suppression of SOD activity in a concentration-dependent manner in green hairy roots. Interestingly, SOD-suppressed root showed three-fold enhanced caffeic acid glucoside accumulation in the soluble fraction as compared to untreated ones. While ascorbate peroxidase activity showed marginal increase in green hairy roots, a decrease in the activities of guaiacol peroxidase and catalase were observed. SDS-PAGE of crude protein profile from green hairy roots showed a distinct band, which was absent in normal hairy roots. MALDI-TOF-MS/MS analysis of the extracted protein confirmed it as the large subunit of RuBisCO. RT-PCR based expression analysis of betaine aldehyde dehydrogenase showed enhanced transcript levels in green hairy roots as compared to normal hairy roots, whereas reverse trends were observed with the transcripts accumulation for phenylalanine ammonia-lyase and chalcone synthase. These findings corroborate with the in vitro BADH activities in hairy roots, and thus indicate an important role of this stress enzyme in combating photo-oxidative stress in green hairy roots upon continuous light exposure. Copyright © 2013 Elsevier GmbH. All rights reserved.

Micropropagation and hairy root culture of Ophiorrhiza alata Craib for camptothecin production.

PubMed

Ya-ut, Pornwilai; Chareonsap, Piyarat; Sukrong, Suchada

2011-12-01

An efficient system was developed for the in vitro micropropagation and hairy root culture of Ophiorrhiza alata Craib for camptothecin (CPT) production. Shoot multiplication on leaf and node explants from germinated seeds of O. alata was successful on half-strength Murashige and Skoog medium supplemented with varying amounts of kinetin and α-naphthaleneacetic acid. Node explants grown in vitro were successfully infected by Agrobacterium rhizogenes TISTR 1450 for the establishment of hairy root culture. The amount of CPT in various parts of O. alata was analyzed by HPLC. The accumulation of CPT in transformed hairy roots was twice that in soil-grown plants (785 ± 52 and 388 ± 32 μg/g dry wt, respectively). In the presence of a polystyrene resin (Diaion HP-20) that absorbed CPT, the CPT content in the culture media increased sevenfold compared with controls (1,036 and 151 μg per 250 ml medium, respectively). These results enable the feasible production of CPT of O. alata by means of a cell culture strategy. These measures can help safeguard the plant from extinction. © Springer Science+Business Media B.V. 2011

Hyperaccumulation of cadmium by hairy roots of Thlaspi caerulescens

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nedelkoska, T.V.; Doran, P.M.

Hairy roots were used to investigate cadmium uptake by Thlaspi caerulescens, a metal hyperaccumulator plant with potential applications in phytoremediation and phytomining. Experiments were carried out in nutrient media under conditions supporting root growth. Accumulation of Cd in short-term (9-h) experiments varied with initial medium pH and increased after treating the roots with H{sup +}-ATPase inhibitor. The highest equilibrium Cd content measured in T. caerulescens roots was 62,800 {micro}g g{sup {minus}1} dry weight, or 6.3% dry weight, at a liquid Cd concentration of 3,710 ppm. Cd levels in live T. caerulescens roots were 1.5- to 1.7-fold those in hairy rootsmore » of nonhyperaccumulator species exposed to the same Cd concentration, but similar to the Cd content of auto-claved T. caerulescens roots. The ability to grow at Cd concentrations of up to 100 ppm clearly distinguished T. caerulescens hairy roots from the nonhyperaccumulators. The specific growth rate of T. caerulescens roots was essentially unaffected by 20 to 50 ppm Cd in the culture medium; in contrast, N. tabacum roots turned dark brown at 20 ppm and growth was negligible. Up to 10,600 {micro}g g{sup {minus}1} dry weight Cd was accumulated by growing T. caerulescens hairy roots. Measurement of Cd levels in while roots and in the cell wall fraction revealed significant differences in the responses of T. caerulescens and N. tabacum roots to 20 ppm Cd. Most metal was transported directly into the symplasm of N. tabacum roots within 3 days of exposure; in contrast, T. caerulescens roots stored virtually all of their Cd in the wall fraction for the first 7 to 10 days. This delay in transmembrane uptake may represent an important defensive strategy against Cd poisoning in T. caerulescens, allowing time for activation of intracellular mechanisms for heavy metal detoxification.« less

Enhanced load-carrying capacity of hairy surfaces floating on water.

PubMed

Xue, Yahui; Yuan, Huijing; Su, Weidong; Shi, Yipeng; Duan, Huiling

2014-05-08

Water repellency of hairy surfaces depends on the geometric arrangement of these hairs and enables different applications in both nature and engineering. We investigate the mechanism and optimization of a hairy surface floating on water to obtain its maximum load-carrying capacity by the free energy and force analyses. It is demonstrated that there is an optimum cylinder spacing, as a result of the compromise between the vertical capillary force and the gravity, so that the hairy surface has both high load-carrying capacity and mechanical stability. Our analysis makes it clear that the setae on water striders' legs or some insects' wings are in such an optimized geometry. Moreover, it is shown that surface hydrophobicity can further increase the capacity of a hairy surface with thick cylinders, while the influence is negligible when the cylinders are thin.

Enhanced load-carrying capacity of hairy surfaces floating on water

PubMed Central

Xue, Yahui; Yuan, Huijing; Su, Weidong; Shi, Yipeng; Duan, Huiling

2014-01-01

Water repellency of hairy surfaces depends on the geometric arrangement of these hairs and enables different applications in both nature and engineering. We investigate the mechanism and optimization of a hairy surface floating on water to obtain its maximum load-carrying capacity by the free energy and force analyses. It is demonstrated that there is an optimum cylinder spacing, as a result of the compromise between the vertical capillary force and the gravity, so that the hairy surface has both high load-carrying capacity and mechanical stability. Our analysis makes it clear that the setae on water striders' legs or some insects' wings are in such an optimized geometry. Moreover, it is shown that surface hydrophobicity can further increase the capacity of a hairy surface with thick cylinders, while the influence is negligible when the cylinders are thin. PMID:24808757

Hairy carbon electrodes studied by cyclic voltammetry and battery discharge testing

NASA Technical Reports Server (NTRS)

Chung, Deborah D. L.; Shui, Xiaoping; Frysz, Christine A.

1993-01-01

Hairy carbon is a new material developed by growing submicron carbon filaments on conventional carbon substrates. Typical substrate materials include carbon black, graphite powder, carbon fibers, and glassy carbon. A catalyst is used to initiate hair growth with carbonaceous gases serving as the carbon source. To study the electrochemical behavior of hairy carbons, cyclic voltammetry (CV) and discharge testing were conducted. In both cases, hairy carbon results surpassed those of the substrate material alone.

Light Requirement for Shoot Regeneration in Horseradish Hairy Roots 1

PubMed Central

Saitou, Tsutomu; Kamada, Hiroshi; Harada, Hiroshi

1992-01-01

Hairy roots of horseradish (Armoracia rusticana) were induced by inoculation with Agrobacterium rhizogenes harboring Ri plasmid and cultured on phytohormone-free Murashige and Skoog medium after eliminating the bacteria. Hairy roots grew vigorously and sometimes formed yellowish calli under dark conditions. On the other hand, growth of hairy roots stopped after several weeks of culture with light, then shoots were regenerated. Frequency of shoot formation from hairy roots increased as the culture period in light lengthened and the light intensity increased. The shoot regeneration was induced by treatment with white or red light, but not with far-red light. Shoot regeneration by red light was inhibited by following treatment with far-red light. Red and far-red light reversibly affected shoot regeneration. Excised roots of nontransformed plants grew quite slowly on phytohormone-free Murashige and Skoog medium and occasionally formed shoots under white light conditions. PMID:16669041

Hairy black holes and the endpoint of AdS4 charged superradiance

NASA Astrophysics Data System (ADS)

Dias, Óscar J. C.; Masachs, Ramon

2017-02-01

We construct hairy black hole solutions that merge with the anti-de Sitter (AdS4) Reissner-Nordström black hole at the onset of superradiance. These hairy black holes have, for a given mass and charge, higher entropy than the corresponding AdS4-Reissner-Nordström black hole. Therefore, they are natural candidates for the endpoint of the charged superradiant instability. On the other hand, hairy black holes never dominate the canonical and grand-canonical ensembles. The zero-horizon radius of the hairy black holes is a soliton (i.e. a boson star under a gauge transformation). We construct our solutions perturbatively, for small mass and charge, so that the properties of hairy black holes can be used to testify and compare with the endpoint of initial value simulations. We further discuss the near-horizon scalar condensation instability which is also present in global AdS4-Reissner-Nordström black holes. We highlight the different nature of the near-horizon and superradiant instabilities and that hairy black holes ultimately exist because of the non-linear instability of AdS.

The BTK Inhibitor Ibrutinib (PCI-32765) Blocks Hairy Cell Leukaemia Survival, Proliferation and BCR Signalling: A New Therapeutic Approach

PubMed Central

Sivina, Mariela; Kreitman, Robert J.; Arons, Evgeny; Ravandi, Farhad; Burger, Jan A.

2014-01-01

B cell receptor (BCR) signalling plays a critical role in the progression of several B-cell malignancies, but its role in hairy cell leukaemia (HCL) is ambiguous. Bruton tyrosine kinase (BTK), a key player in BCR signalling, migration and adhesion, can be targeted with ibrutinib, a selective, irreversible BTK inhibitor. We analysed BTK expression and function in HCL and analysed the effects of ibrutinib on HCL cells. We demonstrated uniform BTK protein expression in HCL cells. Ibrutinib significantly inhibited HCL proliferation and cell cycle progression. Accordingly, ibrutinib also reduced HCL cell survival after BCR triggering with anti-immunoglobulins (A, G, and M) and abrogated the activation of kinases downstream of the BCR (PI3K and MAPK). Ibrutinib also inhibited BCR-dependent secretion of the chemokines CCL3 and CCL4 by HCL cells. Interestingly, ibrutinib inhibited CXCL12-induced signalling, a key pathway for bone marrow homing. Collectively, our data support the clinical development of ibrutinib in patients with HCL. PMID:24697238

The bruton tyrosine kinase inhibitor ibrutinib (PCI-32765) blocks hairy cell leukaemia survival, proliferation and B cell receptor signalling: a new therapeutic approach.

PubMed

Sivina, Mariela; Kreitman, Robert J; Arons, Evgeny; Ravandi, Farhad; Burger, Jan A

2014-07-01

B cell receptor (BCR) signalling plays a critical role in the progression of several B-cell malignancies, but its role in hairy cell leukaemia (HCL) is ambiguous. Bruton tyrosine kinase (BTK), a key player in BCR signalling, as well as B cell migration and adhesion, can be targeted with ibrutinib, a selective, irreversible BTK inhibitor. We analysed BTK expression and function in HCL and analysed the effects of ibrutinib on HCL cells. We demonstrated uniform BTK protein expression in HCL cells. Ibrutinib significantly inhibited HCL proliferation and cell cycle progression. Accordingly, ibrutinib also reduced HCL cell survival after BCR triggering with anti-immunoglobulins and abrogated the activation of kinases downstream of the BCR (PI3K and MAPK). Ibrutinib also inhibited BCR-dependent secretion of the chemokines CCL3 and CCL4 by HCL cells. Interestingly, ibrutinib inhibited also CXCL12-induced signalling, a key pathway for bone marrow homing. Collectively, our data support the clinical development of ibrutinib in patients with HCL. © 2014 John Wiley & Sons Ltd.

Recommendations of the SFH (French Society of Haematology) for the diagnosis, treatment and follow-up of hairy cell leukaemia.

PubMed

Cornet, Edouard; Delmer, Alain; Feugier, Pierre; Garnache-Ottou, Francine; Ghez, David; Leblond, Véronique; Levy, Vincent; Maloisel, Frédéric; Re, Daniel; Zini, Jean-Marc; Troussard, Xavier

2014-12-01

Hairy cell leukaemia (HCL) is a rare haematological malignancy, with approximately 175 new incident cases in France. Diagnosis is based on a careful examination of the blood smear and immunophenotyping of the tumour cells, with a panel of four markers being used specifically to screen for hairy cells (CD11c, CD25, CD103 and CD123). In 2011, the V600E mutation of the BRAF gene in exon 15 was identified in HCL; being present in HCL, it is absent in the variant form of HCL (HCL-v) and in splenic red pulp lymphoma (SRPL), two entities related to HCL. The management of patients with HCL has changed in recent years. A poorer response to purine nucleoside analogues (PNAs) is observed in patients with more marked leukocytosis, bulky splenomegaly, an unmutated immunoglobulin variable heavy chain (IgVH) gene profile, use of VH4-34 or with TP53 mutations. We present the recommendations of a group of 11 experts belonging to a number of French hospitals. This group met in November 2013 to examine the criteria for managing patients with HCL. The ideas and proposals of the group are based on a critical analysis of the recommendations already published in the literature and on an analysis of the practices of clinical haematology departments with experience in managing these patients. The first-line treatment uses purine analogues: cladribine or pentostatin. The role of BRAF inhibitors, whether or not combined with MEK inhibitors, is discussed. The panel of French experts proposed recommendations to manage patients with HCL, which can be used in a daily practice.

Aspects of hairy black holes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Anabalón, Andrés, E-mail: andres.anabalon-at@uai.cl; Astefanesei, Dumitru

We review the existence of exact hairy black holes in asymptotically flat, anti-de Sitter and de Sitter space-times. We briefly discuss the issue of stability and the charging of the black holes with a Maxwell field.

Psoralen production in hairy roots and adventitious roots cultures of Psoralea coryfolia.

PubMed

Baskaran, P; Jayabalan, N

2009-07-01

Psoralea corylifolia is an endangered plant producing various compounds of medical importance. Adventitious roots and hairy roots were induced in cultures prepared from hypocotyl explants. Psoralen content was evaluated in both root types grown either in suspension cultures or on agar solidified medium. Psoralen content was approximately 3 mg g(-1) DW in suspension grown hairy roots being higher than in solid grown hairy roots and in solid and suspension-grown adventitious roots.

Redirection of metabolite biosynthesis from hydroxybenzoates to volatile terpenoids in green hairy roots of Daucus carota.

PubMed

Mukherjee, Chiranjit; Samanta, Tanmoy; Mitra, Adinpunya

2016-02-01

A metabolic shift in green hairy root cultures of carrot from phenylpropanoid/benzenoid biosynthesis toward volatile isoprenoids was observed when compared with the metabolite profile of normal hairy root cultures. Hairy roots cultures of Daucus carota turned green under continuous illumination, while the content of the major phenolic compound p-hydroxybenzoic acid (p-HBA) was reduced to half as compared to normal hairy roots cultured in darkness. p-Hydroxybenzaldehyde dehydrogenase (HBD) activity was suppressed in the green hairy roots. However, comparative volatile analysis of 14-day-old green hairy roots revealed higher monoterpene and sesquiterpene contents than found in normal hairy roots. Methyl salicylate content was higher in normal hairy roots than in green ones. Application of clomazone, an inhibitor of 1-deoxy-D-xylulose 5-phosphate synthase (DXS), reduced the amount of total monoterpenes and sesquiterpenes in green hairy roots compared to normal hairy roots. However, methyl salicylate content was enhanced in both green and normal hairy roots treated with clomazone as compared to their respective controls. Because methyl-erythritol 4-phosphate (MEP) and phenylpropanoid pathways, respectively, contribute to the formation of monoterpenes and phenolic acids biosynthesis, the activities of enzymes regulating those pathways were measured in terms of their in vitro activities, in both green and normal hairy root cultures. These key enzymes were 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR), an early regulatory enzyme of the MEP pathway, pyruvate kinase (PK), an enzyme of primary metabolism related to the MEP pathway, shikimate dehydrogenase (SKDH) which is involved in biosynthesis of aromatic amino acids, and phenylalanine ammonia-lyase (PAL) that catalyzes the first step of phenylpropanoid biosynthesis. Activities of DXR and PK were higher in green hairy roots as compared to normal ones, whereas the opposite trend was observed for SKDH and PAL

Hairy root biotechnology--indicative timeline to understand missing links and future outlook.

PubMed

Mehrotra, Shakti; Srivastava, Vikas; Ur Rahman, Laiq; Kukreja, A K

2015-09-01

Agrobacterium rhizogenes-mediated hairy roots (HR) were developed in the laboratory to mimic the natural phenomenon of bacterial gene transfer and occurrence of disease syndrome. The timeline analysis revealed that during 90 s, the research expanded to the hairy root-based secondary metabolite production and different yield enhancement strategies like media optimization, up-scaling, metabolic engineering etc. An outlook indicates that much emphasis has been given to the strategies that are helpful in making this technology more practical in terms of high productivity at low cost. However, a sequential analysis of literature shows that this technique is upgraded to a biotechnology platform where different intra- and interdisciplinary work areas were established, progressed, and diverged to provide scientific benefits of various hairy root-based applications like phytoremediation, molecular farming, biotransformation, etc. In the present scenario, this biotechnology research platform includes (a) elemental research like hairy root-mediated secondary metabolite production coupled with productivity enhancement strategies and (b) HR-based functional research. The latter comprised of hairy root-based applied aspects such as generation of agro-economical traits in plants, production of high value as well as less hazardous molecules through biotransformation/farming and remediation, respectively. This review presents an indicative timeline portrayal of hairy root research reflected by a chronology of research outputs. The timeline also reveals a progressive trend in the state-of-art global advances in hairy root biotechnology. Furthermore, the review also discusses ideas to explore missing links and to deal with the challenges in future progression and prospects of research in all related fields of this important area of plant biotechnology.

Gentiana dinarica Beck hairy root cultures and evaluation of factors affecting growth and xanthone production

USDA-ARS?s Scientific Manuscript database

The induction and establishment of hairy root cultures of Gentiana dinarica using two strains of Agrobacterium rhizogenes (A4M70GUS and 15834/PI) is reported for the first time. Hairy roots were formed from the shoots 25 days after inoculation, and strain 15834/PI had higher induction rate of hairy ...

Rotating hairy black holes in arbitrary dimensions

NASA Astrophysics Data System (ADS)

Erices, Cristián; Martínez, Cristián

2018-01-01

A class of exact rotating black hole solutions of gravity nonminimally coupled to a self-interacting scalar field in arbitrary dimensions is presented. These spacetimes are asymptotically locally anti-de Sitter manifolds and have a Ricci-flat event horizon hiding a curvature singularity at the origin. The scalar field is real and regular everywhere, and its effective mass, coming from the nonminimal coupling with the scalar curvature, saturates the Breitenlohner-Freedman bound for the corresponding spacetime dimension. The rotating black hole is obtained by applying an improper coordinate transformation to the static one. Although both spacetimes are locally equivalent, they are globally different, as it is confirmed by the nonvanishing angular momentum of the rotating black hole. It is found that the mass is bounded from below by the angular momentum, in agreement with the existence of an event horizon. The thermodynamical analysis is carried out in the grand canonical ensemble. The first law is satisfied, and a Smarr formula is exhibited. The thermodynamical local stability of the rotating hairy black holes is established from their Gibbs free energy. However, the global stability analysis establishes that the vacuum spacetime is always preferred over the hairy black hole. Thus, the hairy black hole is likely to decay into the vacuum one for any temperature.

Production and analysis of organic acids in hairy-root cultures of Isatis indigotica Fort. (indigo woad).

PubMed

Xu, Tiefeng; Zhang, Lei; Sun, Xiaofen; Zhang, Hanming; Tang, Kexuan

2004-02-01

Hairy roots were induced from both cotyledon and hypocotyl explants of Isatis indigotica Fort. (indigo woad) through transformation with Agrobaterium rhizogenes strain A4, R1601 and ATCC15834. The results showed that the cotyledons were the preferred explants to hypocotyls and A4 was the most suitable A. rhizogenes strain for the transformation and induction of hairy roots of I. indigotica. High-voltage paper electrophoresis (HVPE) analysis demonstrated the production of mannopine in hairy roots and confirmed the successful transfer of Ri T-DNA (root-inducing transferred DNA) of A. rhizogenes into the I. indigotica genome. Five organic acids, namely CPQ [3-(2-carboxyphenol)-4(3 H )-quinazolinone], syringic acid, salicylic acid, benzoic acid and 2-aminobenzoic acid, which were considered as main antiviral components of I. indigotica, were detected in natural roots, hairy roots and liquid media with high-performance capillary electrophoresis. The results showed CPQ production in hairy roots was significantly higher than that in natural roots. Our results also revealed that all the five organic acids could be excreted from hairy roots into liquid media, and the concentrations of organic acids in the liquid media paralleled those in hairy roots. The hairy roots of I. indigotica grew fast and showed an S-shaped growth curve that reached its apex on the day 24 of culture with a 20-fold increase in fresh weight compared with the starting inoculums. The accumulation of the two organic acids CPQ and syringic acid in liquid media paralleled the growth of hairy roots. MS [Murashige, T. and Skoog, F. (1962) Physiol. Plant. 15, 473-497] medium or half-strength MS medium supplemented with 30 g/l maltose was found to be best for hairy-root culture and accumulation of CPQ.

Audit of compliance with the British Committee for Standards in Haematology (BCSH) revised guidelines for the diagnosis and assessment of treatment response of hairy cell leukemia in University Hospital Galway.

PubMed

Velazquez-Kennedy, K; Crowe, C; Craven, B; Walsh, J; Prendergast, C; Krawczyk, J

2017-05-01

Hairy cell leukemia (HCL) is an uncommon B cell lymphoproliferative disorder. The object of the present audit was to assess whether the investigation and management of HCL in University College Hospital Galway (UCHG) complies with the British Committee for Standards in Haematology (BCSH) guidelines. Following a review of the records in our Haematology Department, 18 cases of HCL were identified between January 2006 and October 2014. Blood film examination had been performed in all cases. Flow cytometry of liquid material had been undertaken in 89 % (n = 16) of cases, of which only 31 % (n = 5) included all four hairy cell panel markers (CD11c, CD25, CD103, CD123). Although all initial trephine biopsies included CD20, none analyzed DBA44. Only 65 % (n = 11) of treated patients had a post-treatment bone marrow biopsy preformed. This audit highlights areas of improvement in the diagnosis and management of HCL in UCHG, which do not currently adhere to the BCSH recommendations.

Lignan enhancement in hairy root cultures of Linum album using coniferaldehyde and methylenedioxycinnamic acid.

PubMed

Ahmadian Chashmi, Najmeh; Sharifi, Mohsen; Behmanesh, Mehrdad

2016-07-03

Feeding experiments with hairy root cultures of Linum album have established that the extracellular coniferaldehyde is a good precursor for production of two lignans: lariciresinol (LARI) and pinoresinol (PINO). The accumulation of the LARI, PINO, and podophyllotoxin (PTOX) in hairy roots were enhanced about 14.8-, 8.7-, and 1.5-fold (107.61, 8.7 and 6.42 µg g(-1) Fresh Wight), respectively, by the addition of coniferaldehyde (2 mM) to the culture media (after 24 hr). This result was correlated with an increase pinoresinol/lariciresinol reductase (PLR) expression gene and cinnamyl alcohol dehydrogenase (CAD) activity in the fed hairy roots. Adding 3,4-(methylendioxy)cinnamic acid (MDCA) precursor did not influence on the lignans accumulation, but the lignin content of the hairy roots was increased. Moreover, the expression genes of phenylalanine ammonialyase (PAL), CAD, and cinnamoyl-CoA reductase (CCR) were influenced after feeding hairy roots with MDCA.

Sennosides A and B production by hairy roots of Senna alata (L.) Roxb.

PubMed

Putalun, Waraporn; Pimmeuangkao, Suwat; De-Eknamkul, Wanchai; Tanaka, Hiroyuki; Shoyama, Yukihiro

2006-01-01

Hairy roots of Senna alata transformed with Agrobacterium rhizogenes, strain ATCC 15834 were induced and grown in half-strength Murashige and Skoog (MS) medium. Effects of sucrose contents and hormones on the growth and sennosides A, B production were investigated. Hairy roots cultured on hormone-free half-strength MS medium containing 5% sucrose under dark condition mostly stimulated the growth of hairy roots and increased the content of sennosides A and B yielding (169 +/- 4) and (34 +/- 3) microg g(-1) dry wt, respectively.

Metabolomic Analysis and Phenylpropanoid Biosynthesis in Hairy Root Culture of Tartary Buckwheat Cultivars

PubMed Central

Li, Xiaohua; Bok Kim, Yeon; Romij Uddin, Md; Kim, Sun Ju; Suzuki, Tatsuro; Park, Nam Il; Park, Sang Un

2013-01-01

Buckwheat, Fagopyrum tataricum Gaertn., is an important medicinal plant, which contains several phenolic compounds, including one of the highest content of rutin, a phenolic compound with anti-inflammatory properties. An experiment was conducted to investigate the level of expression of various genes in the phenylpropanoid biosynthetic pathway to analyze in vitro production of anthocyanin and phenolic compounds from hairy root cultures derived from 2 cultivars of tartary buckwheat (Hokkai T8 and T10). A total of 47 metabolites were identified by gas chromatography–time-of-flight mass spectrometry (GC-TOFMS) and subjected to principal component analysis (PCA) in order to fully distinguish between Hokkai T8 and T10 hairy roots. The expression levels of phenylpropanoid biosynthetic pathway genes, through qRT-PCR, showed higher expression for almost all the genes in T10 than T8 hairy root except for FtF3’H-2 and FtFLS-2. Rutin, quercetin, gallic acid, caffeic acid, ferulic acid, 4-hydroxybenzoic acid, and 2 anthocyanin compounds were identified in Hokkai T8 and T10 hairy roots. The concentration of rutin and anthocyanin in Hokkai T10 hairy roots of tartary buckwheat was several-fold higher compared with that obtained from Hokkai T8 hairy root. This study provides useful information on the molecular and physiological dynamic processes that are correlated with phenylpropanoid biosynthetic gene expression and phenolic compound content in F. tataricum species. PMID:23799007

Scale invariant hairy black holes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Banados, Maximo; Theisen, Stefan

Scalar fields coupled to three-dimensional gravity are considered. We uncover a scaling symmetry present in the black hole reduced action, and use it to prove a Smarr formula valid for any potential. We also prove that nonrotating hairy black holes exists only for positive total energy. The extension to higher dimensions is also considered.

Hairy black holes in scalar extended massive gravity

NASA Astrophysics Data System (ADS)

Tolley, Andrew J.; Wu, De-Jun; Zhou, Shuang-Yong

2015-12-01

We construct static, spherically symmetric black hole solutions in scalar extended ghost-free massive gravity and show the existence of hairy black holes in this class of extension. While the existence seems to be a generic feature, we focus on the simplest models of this extension and find that asymptotically flat hairy black holes can exist without fine-tuning the theory parameters, unlike the bi-gravity extension, where asymptotical flatness requires fine-tuning in the parameter space. Like the bi-gravity extension, we are unable to obtain asymptotically dS regular black holes in the simplest models considered, but it is possible to obtain asymptotically AdS black holes.

Study of artemisinin and sugar accumulation in Artemisia vulgaris and Artemisia dracunculus "hairy" root cultures.

PubMed

Drobot, Kateryna O; Matvieieva, Nadiia A; Ostapchuk, Andriy M; Kharkhota, Maxim A; Duplij, Volodymyr P

2017-09-14

We studied the effect of genetic transformation on biologically active compound (artemisinin and its co-products (ART) as well as sugars) accumulation in Artemisia vulgaris and Artemisia dracunculus "hairy" root cultures. Glucose, fructose, sucrose, and mannitol were accumulated in A. vulgaris and A. dracunculus "hairy" root lines. Genetic transformation has led in some cases to the sugar content increasing or appearing of nonrelevant for the control plant carbohydrates. Sucrose content was 1.6 times higher in A. vulgaris "hairy" root lines. Fructose content was found to be 3.4 times higher in A. dracunculus "hairy" root cultures than in the control roots. The accumulation of mannitol was a special feature of the leaves of A. vulgaris and A. dracunculus control roots. A. vulgaris "hairy" root lines differed also in ART accumulation level. The increase of ART content up to 1.02 mg/g DW in comparison with the nontransformed roots (up to 0.687 mg/g DW) was observed. Thus, Agrobacterium rhizogenes-mediated genetic transformation can be used for obtaining of A. vulgaris and A. dracunculus "hairy" root culture produced ART and sugars in a higher amount than mother plants.

Hairy Root as a Model System for Undergraduate Laboratory Curriculum and Research

ERIC Educational Resources Information Center

Keyes, Carol A.; Subramanian, Senthil; Yu, Oliver

2009-01-01

Hairy root transformation has been widely adapted in plant laboratories to rapidly generate transgenic roots for biochemical and molecular analysis. We present hairy root transformations as a versatile and adaptable model system for a wide variety of undergraduate laboratory courses and research. This technique is easy, efficient, and fast making…

Production of the biopesticide azadirachtin by hairy root cultivation of Azadirachta indica in liquid-phase bioreactors.

PubMed

Srivastava, Smita; Srivastava, Ashok K

2013-11-01

Batch cultivation of Azadirachta indica hairy roots was carried out in different liquid-phase bioreactor configurations (stirred-tank, bubble column, bubble column with polypropylene basket, and polyurethane foam disc as root supports) to investigate possible scale-up of the A. indica hairy root culture for in vitro production of the biopesticide azadirachtin. The hairy roots failed to grow in the conventional bioreactor designs (stirred tank and bubble column). However, modified bubble column reactor (with polyurethane foam as root support) configuration facilitated high-density culture of A. indica hairy roots with a biomass production of 9.2 g l(-1)dry weight and azadirachtin yield of 3.2 mg g(-1) leading to a volumetric productivity of azadirachtin as 1.14 mg l(-1) day(-1). The antifeedant activity in the hairy roots was also evaluated by no choice feeding tests with known concentrations of the hairy root powder and its solvent extract separately on the desert locust Schistocerca gregaria. The hairy root powder and its solvent extract demonstrated a high level of antifeedant activity (with an antifeedant index of 97 % at a concentration of 2 % w/v and 83 % at a concentration of 0.05 % (w/v), respectively, in ethanol).

Tunable Pickering Emulsions with Environmentally Responsive Hairy Silica Nanoparticles.

PubMed

Liu, Min; Chen, Xiaoli; Yang, Zongpeng; Xu, Zhou; Hong, Liangzhi; Ngai, To

2016-11-30

Surface modification of the nanoparticles using surface anchoring of amphiphilic polymers offers considerable scope for the design of a wide range of brush-coated hybrid nanoparticles with tunable surface wettability that may serve as new class of efficient Pickering emulsifiers. In the present study, we prepared mixed polymer brush-coated nanoparticles by grafting ABC miktoarm star terpolymers consisting of poly(ethylene glycol), polystyrene, and poly[(3-triisopropyloxysilyl)propyl methacrylate] (μ-PEG-b-PS-b-PIPSMA) on the surface of silica nanoparticles. The wettability of the as-prepared nanoparticles can be precisely tuned by a change of solvent or host-guest complexation. 1 H NMR result confirmed that such wettability change is due to the reorganization of the polymer chain at the grafted layer. We show that this behavior can be used for stabilization and switching between water-in-oil (W/O) and oil-in-water (O/W) emulsions. For hairy particles initially dispersed in oil, W/O emulsions were always obtained with collapsed PEG chains and mobile PS chains at the grafted layer. However, initially dispersing the hairy particles in water resulted in O/W emulsions with collapsed PS chains and mobile PEG chains. When a good solvent for both PS and PEG blocks such as toluene was used, W/O emulsions were always obtained no matter where the hairy particles were dispersed. The wettability of the mixed polymer brush-coated silica particles can also be tuned by host-guest complexation between PEG block and α-CD. More importantly, our result showed that surprisingly the resultant mixed brush-coated hairy nanoparticles can be employed for the one-step production of O/W/O multiple emulsions that are not attainable from conventional Pickering emulsifiers. The functionalized hairy silica nanoparticles at the oil-water interface can be further linked together utilizing poly(acrylic acid) as the reversible linker to form supramolecular colloidosomes, which show p

Glucosinolate biosynthesis in hairy root cultures of broccoli (Brassica oleracea var. italica).

PubMed

Kim, Sun-Ju; Park, Woo Tae; Uddin, Md Romij; Kim, Yeon Bok; Nam, Sang-Yong; Jho, Kwang Hyun; Park, Sang Un

2013-02-01

Here we present previously unreported glucosinolate production by hairy root cultures of broccoli (B. oleracea var. italica). Growth media greatly influenced the growth and glucosinolate content of hairy root cultures of broccoli. Seven glucosinolates, glucoraphanin, gluconapin, glucoerucin, glucobrassicin, 4-methoxyglucobrassicin, gluconasturtiin, and neoglucobrassicin, were identified by analysis of the broccoli hairy root cultures. Both half and full strength B5 and SH media enabled the highest accumulation of glucosinolates. In most cases, the levels of glucosinolates were higher in SH and BS media. Among the 7 glucosinolates, the accumulation of neoglucobrassicin was very high, irrespective of growth medium. The neoglucobrassicin content was 7.4-fold higher in SH medium than 1/2 MS, in which its level was the lowest. The 1/2 B5 medium supported the production of the highest amounts of glucobrassicin and 4-methoxyglucobrassicin, the levels for which were 36.2- and 7.9- fold higher, respectively, than their lowest content in 1/2 MS medium. The 1/2 SH medium enabled the highest accumulation of glucoraphanin and gluconapin in the broccoli hairy root cultures, whose levels were 1.8- and 4.6-fold higher, respectively, than their lowest content in 1/2 MS medium. Our results suggest that hairy root cultures of broccoli could be a valuable alternative approach for the production of glucosinolate compounds.

Abrasion properties of self-suspended hairy titanium dioxide nanomaterials

NASA Astrophysics Data System (ADS)

Zhang, Jiao-xia; Liu, Si; Yan, Chao; Wang, Xiao-jing; Wang, Lei; Yu, Ya-ming; Li, Shi-yun

2017-11-01

Considering the excellent solubility of pyrrolidone ring organic compounds, the synthesized N-(trimethoxysilyl) propyl- N-methyl-2-pyrrolidone chlorides was tethered onto titanium dioxide (TiO2) nanoparticles to improve dispersion of TiO2, and then polyethylene oxide (PEO) oligomer through ion exchange embraced the tethered TiO2 to obtain a novel self-suspended hairy TiO2 nanomaterials without any solvent. A variety of techniques were carried out to illustrate the structure and properties of the self-suspended hairy TiO2 nanomaterials. It was found that TiO2 nanoparticles embody monodispersity in the hybrid system though the "false reunion" phenomenon occurring due to nonpermanent weak physical cross-linking. Remarkably, self-suspended hairy TiO2 nanomaterials exhibit lower viscosity, facilitating maneuverable and outstanding antifriction and wear resistance properties, due to the synergistic lubricating effect between spontaneously forming lubricating film and nano-lubrication of TiO2 cores, overcoming the deficiency of both solid and liquid lubricants. This make them promising candidates for the micro-electromechanic/nano-electromechanic systems (MEMS/NEMS).

Menthol and geraniol biotransformation and glycosylation capacity of Levisticum officinale hairy roots.

PubMed

Nunes, Inês S; Faria, Jorge M S; Figueiredo, A Cristina; Pedro, Luis G; Trindade, Helena; Barroso, José G

2009-03-01

The biotransformation capacity of Levisticum officinale W.D.J. Koch hairy root cultures was studied by evaluating the effect of the addition of 25 mg/L menthol or geraniol on morphology, growth, and volatiles production. L. officinale hairy root cultures were maintained for 7 weeks in SH medium, in darkness at 24 degrees C and 80 r.p.m., and the substrates were added 15 days after inoculation. Growth was evaluated by measuring fresh and dry weight and by using the dissimilation method. Volatiles composition was analyzed by GC and GC-MS. Hairy roots morphology and growth were not influenced by substrate addition. No new volatiles were detected after menthol addition and, as was also the case with the control cultures, volatiles of these hairy roots were dominated by (Z)-falcarinol (1-45%), N-octanal (3-8%), palmitic acid (3-10%), and (Z)-ligustilide (2-9%). The addition of geraniol induced the production of six new volatiles: nerol/citronellol/neral (traces-15%), alpha-terpineol (0.2-3%), linalool (0.1-1.2%), and geranyl acetate (traces-2%). The relative amounts of the substrates and some of their biotransformation products decreased during the course of the experiment. Following the addition of beta-glycosidase to the remaining distillation water, analysis of the extracted volatiles showed that lovage hairy roots were able to convert both substrates and their biotransformation products into glycosidic forms. GC:gas chromatography GC-MS:gas chromatography-mass spectrometry SH:Schenk and Hildebrandt (1972) culture medium.

Gels from soft hairy nanoparticles in polymeric matrices

NASA Astrophysics Data System (ADS)

Vlassopoulos, Dimitris

2013-03-01

Hairy particles represent a huge class of soft colloids with tunable interactions and properties. Advances in synthetic chemistry have enabled obtaining well-characterized such systems for specific needs. In this talk we present two model hairy soft particles with diameters of the order of tens of nanometers, star polymers and polymerically grafted spherical particles. In particular, we discuss design strategies for dispersing them in polymeric matrices and eventually creating and breaking gels. Control parameters are the matrix molar mass, the grafting density (or functionality) and the size of the grafts (or arms). The linear viscoelastic properties and slow time evolution of the gels are examined in view of the existing knowledge from colloidal gels consisting of micron-sized particles, and compared. In the case of stars we start from a concentrated glassy suspension in molecular solvent and add homopolymer at increasing concentration, and as a result of the induced osmotic pressure the stars shrink and a depletion gel is formed. For the grafted colloidal particles, they are added at low concentration to a polymer matrix, and it has been shown that under certain conditions the anisotropy of interactions gives rise to network formation. We then focus on the nonlinear rheological response and in particular the effect of shear flow in inducing a solid to liquid transition. Our studies show that the yielding process is gradual and shares many common features with that of flocculated colloidal suspensions, irrespectively of the shape of the building block of the gel. Whereas shear can melt such a gel, it cannot break it into its constituent blocks and hence fully disperse the hairy nanoparticles. On the other hand, the hairy particles are intrinsically hybrid. We show how this important feature is reflected on the heating of the gels. In that case, the mismatch of thermal expansion coefficients of core and shell appears to play a role on the particle response as it

Scalar hairy black holes and scalarons in the isolated horizons formalism

DOE Office of Scientific and Technical Information (OSTI.GOV)

Corichi, Alejandro; Instituto de Matematicas, Universidad Nacional Autonoma de Mexico, A. Postal 61-3, Morelia, Michoacan, 58090; Nucamendi, Ulises

The Isolated Horizons (IH) formalism, together with a simple phenomenological model for colored black holes has been used to predict nontrivial formulas that relate the ADM mass of the solitons and hairy Black Holes of Gravity-Matter system on the one hand, and several horizon properties of the black holes in the other. In this article, the IH formalism is tested numerically for spherically symmetric solutions to an Einstein-Higgs system where hairy black holes were recently found to exist. It is shown that the mass formulas still hold and that, by appropriately extending the current model, one can account for themore » behavior of the horizon properties of these new solutions. An empirical formula that approximates the ADM mass of hairy solutions is put forward, and some of its properties are analyzed.« less

Eucalyptus hairy roots, a fast, efficient and versatile tool to explore function and expression of genes involved in wood formation.

PubMed

Plasencia, Anna; Soler, Marçal; Dupas, Annabelle; Ladouce, Nathalie; Silva-Martins, Guilherme; Martinez, Yves; Lapierre, Catherine; Franche, Claudine; Truchet, Isabelle; Grima-Pettenati, Jacqueline

2016-06-01

Eucalyptus are of tremendous economic importance being the most planted hardwoods worldwide for pulp and paper, timber and bioenergy. The recent release of the Eucalyptus grandis genome sequence pointed out many new candidate genes potentially involved in secondary growth, wood formation or lineage-specific biosynthetic pathways. Their functional characterization is, however, hindered by the tedious, time-consuming and inefficient transformation systems available hitherto for eucalypts. To overcome this limitation, we developed a fast, reliable and efficient protocol to obtain and easily detect co-transformed E. grandis hairy roots using fluorescent markers, with an average efficiency of 62%. We set up conditions both to cultivate excised roots in vitro and to harden composite plants and verified that hairy root morphology and vascular system anatomy were similar to wild-type ones. We further demonstrated that co-transformed hairy roots are suitable for medium-throughput functional studies enabling, for instance, protein subcellular localization, gene expression patterns through RT-qPCR and promoter expression, as well as the modulation of endogenous gene expression. Down-regulation of the Eucalyptus cinnamoyl-CoA reductase1 (EgCCR1) gene, encoding a key enzyme in lignin biosynthesis, led to transgenic roots with reduced lignin levels and thinner cell walls. This gene was used as a proof of concept to demonstrate that the function of genes involved in secondary cell wall biosynthesis and wood formation can be elucidated in transgenic hairy roots using histochemical, transcriptomic and biochemical approaches. The method described here is timely because it will accelerate gene mining of the genome for both basic research and industry purposes. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

The Relationship Between Endogenous β-Glucuronidase Activity and Biologically Active Flavones-Aglycone Contents in Hairy Roots of Baikal Skullcap.

PubMed

Dikaya, Varvara S; Solovyeva, Aleksandra I; Sidorov, Roman A; Solovyev, Pavel A; Stepanova, Anna Yu

2018-02-01

Here, we examine the relationship between contents of principal flavones in hairy roots of Scutellaria baicalensis with the activity of the β-glucuronidase (sGUS) enzyme during a culturing cycle. Using RP-HPLC, we show that the highest contents of aglycones, baicalin and wogonin is observed at the growth days 8, 14, and 71 and reach 45, 41, and 62% (based on the total weight of hairy roots of the Baikal skullcap), correspondingly. Their accumulation is accompanied by increase of the sGUS activity, which we determined fluorometrically. Moreover, the enzyme activity is characterized by significant and reasonable correlation only with the wogonin contents. Our results confirm a significant role of sGUS at the final steps of the metabolism in root-specific flavones of Baikal skullcap and suggest how one can optimize the conditions of culturing the hairy roots for biotechnological production of individual flavonoids. For example, at the culturing day 71 wogonin constituted over 80% of all flavones extracted from cells. © 2018 Wiley-VHCA AG, Zurich, Switzerland.

Exposure to pesticides as risk factor for non-Hodgkin's lymphoma and hairy cell leukemia: pooled analysis of two Swedish case-control studies.

PubMed

Hardell, Lennart; Eriksson, Mikael; Nordstrom, Marie

2002-05-01

Increased risk for non-Hodgkin's lymphoma (NHL) following exposure to certain pesticides has previously been reported. To further elucidate the importance of phenoxyacetic acids and other pesticides in the etiology of NHL a pooled analysis was performed on two case-control studies, one on NHL and another on hairy cell leukemia (HCL), a rare subtype of NHL. The studies were population based with cases identified from cancer registry and controls from population registry. Data assessment was ascertained by questionnaires supplemented over the telephone by specially trained interviewers. The pooled analysis of NHL and HCL was based on 515 cases and 1141 controls. Increased risks in univariate analysis were found for subjects exposed to herbicides (OR 1.75, CI 95% 1.26-2.42), insecticides (OR 1.43, CI 95% 1.08-1.87), fungicides (OR 3.11, CI 95% 1.56-6.27) and impregnating agents (OR 1.48, CI 95% 1.11-1.96). Among herbicides, significant associations were found for glyphosate (OR 3.04, CI 95% 1.08-8.52) and 4-chloro-2-methyl phenoxyacetic acid (MCPA) (OR 2.62, CI 95% 1.40-4.88). For several categories of pesticides the highest risk was found for exposure during the latest decades before diagnosis. However, in multivariate analyses the only significantly increased risk was for a heterogeneous category of other herbicides than above.

Fluid and particle transport of a hairy structure

NASA Astrophysics Data System (ADS)

Lee, Hongki; Lahooti, Mohsen; Kim, Daegyoum; Jung, Seyeong

2017-11-01

Hairy appendages of animals are used to capture particles, sense surrounding flow, and generate propulsive force. Due to the small size of the hairy structures, their hydrodynamics have been studied mostly in very low Reynolds number. In this work, in a broad range of Reynolds number, O(1) - O(100), flow structure and inertial particle dynamics around an array of two-dimensional cylinders are investigated numerically by using an immersed boundary method. Given flow fields, Maxey-Riley equation is adopted to examine particle dynamics. Here, we discuss the effects of Reynolds number, density ratio of inertial particles and fluid, and distance between cylinders on particle behaviors around a moving structure. In addition, drift volume of inertial particles is correlated with the model parameters.

Blue light decreases tanshinone IIA content in Salvia miltiorrhiza hairy roots via genes regulation.

PubMed

Chen, Ing-Gin J; Lee, Meng-Shiou; Lin, Ming-Kuem; Ko, Chia-Yun; Chang, Wen-Te

2018-06-01

The effect of light-emitting diodes (LEDs) on the production of secondary metabolites in medicinal plants and hairy roots is receiving much attention. The roots and rhizomes of the traditional Chinese medicinal plant Salvia miltiorrhiza Bunge are widely used for treating cardiovascular and cerebrovascular diseases. The main components are liposoluble tanshinones and hydrophilic phenolic acids. Moreover, hairy root culture of S. miltiorrhiza has been used in research of valuable plant-derived secondary metabolites. In this study, we examined the effect of LEDs with different combinations of wavelengths on the content of the main components in hairy roots of S. miltiorrhiza. Tanshinone IIA (TSIIA) content in hairy roots was significantly decreased with all light treatments containing blue light by >60% and was 9 times lower with LED treatment duration changed from 1 week to 3 weeks. HMGR, DXS2, DXR, GGPPS, CPS and CYP76AH1 genes involved in the tanshinone biosynthesis pathway were downregulated by blue light. Furthermore, light quality treatments have different effect on the accumulation of phenolic acids in hairy roots of S. miltiorrhiza. The light treatments 6R3B, 6B3IR, 7RGB and 2R6BUV for 3 weeks could increase rosmarinic acid (RA) content slightly but not salvianolic acid B (SAB) content. Different secondary metabolite contents could be regulated by different wavelength combinations of LEDs. Blue light could reduce TSIIA content in hairy roots of S. miltiorrhiza via gene regulation. Copyright © 2018. Published by Elsevier B.V.

Analysis of hairy root culture of Rauvolfia serpentina using direct analysis in real time mass spectrometric technique.

PubMed

Madhusudanan, K P; Banerjee, Suchitra; Khanuja, Suman P S; Chattopadhyay, Sunil K

2008-06-01

The applicability of a new mass spectrometric technique, DART (direct analysis in real time) has been studied in the analysis of the hairy root culture of Rauvolfia serpentina. The intact hairy roots were analyzed by holding them in the gap between the DART source and the mass spectrometer for measurements. Two nitrogen-containing compounds, vomilenine and reserpine, were characterized from the analysis of the hairy roots almost instantaneously. The confirmation of the structures of the identified compounds was made through their accurate molecular formula determinations. This is the first report of the application of DART technique for the characterization of compounds that are expressed in the hairy root cultures of Rauvolfia serpentina. Moreover, this also constitutes the first report of expression of reserpine in the hairy root culture of Rauvolfia serpentina. Copyright (c) 2008 John Wiley & Sons, Ltd.

Effect of elicitors and precursors on azadirachtin production in hairy root culture of Azadirachta indica.

PubMed

Srivastava, Smita; Srivastava, A K

2014-02-01

The present study involved strategies for enhancement in in vitro azadirachtin (commercially used biopesticide) production by hairy root cultivation of Azadirachta indica. Improvement in the azadirachtin production via triggering its biosynthetic pathway in plant cells was carried out by the exogenous addition of precursors and elicitors in the growth medium. Among the different abiotic stress inducers (Ag(+), Hg(+2), Co(+2), Cu(+2)) and signal molecules (methyl jasmonate and salicylic acid) tested, salicylic acid at 15 mg l(-1) of concentration was found to enhance the azadirachtin yield in the hairy roots to the maximum (up to 4.95 mg g(-1)). Similarly, among the different biotic elicitors tested (filter-sterilized fungal culture filtrates of Phoma herbarium, Alternaria alternata, Myrothecium sp., Fusarium solani, Curvularia lunata, and Sclerotium rolfsii; yeast extract; and yeast extract carbohydrate fraction), addition of filter-sterilized fungal culture filtrate of C. lunata (1 % v/v) resulted in maximum azadirachtin yield enhancement in hairy root biomass (up to 7.1 mg g(-1)) with respect to the control (3.3 mg g(-1)). Among all the biosynthetic precursors studied (sodium acetate, cholesterol, squalene, isopentynyl pyrophosphate, mavalonic acid lactone, and geranyl pyrophosphate), the overall azadirachtin production (70.42 mg l(-1) in 25 days) was found to be the highest with cholesterol (50 mg l(-1)) addition as an indirect precursor in the medium.

Genetic Transformation and Hairy Root Induction Enhance the Antioxidant Potential of Lactuca serriola L.

PubMed

El-Esawi, Mohamed A; Elkelish, Amr; Elansary, Hosam O; Ali, Hayssam M; Elshikh, Mohamed; Witczak, Jacques; Ahmad, Margaret

2017-01-01

Lactuca serriola L. is a herbaceous species, used for human nutrition and medicinal purposes. The high antioxidant capacity of L. serriola indicates the possibility of enhancing its edible and health potential by increasing the flavonoid and phenolic contents. The present study aimed at enhancing the production of phenolics and flavonoids by hairy root cultures in Lactuca serriola transformed with Agrobacterium rhizogenes strain AR15834 harbouring the rolB gene. The genetic transformation of rolB in transformed roots was validated, and rolB expression level was evaluated using real-time qPCR analysis. Expression levels of flavonoid biosynthesis genes (CHI, PAL, FLS, and CHS) were assessed in the hairy and nontransformed roots. Results showed higher expression levels in the transgenic roots than in the nontransformed ones ( p < 0.01). Transgenic hairy roots exhibited a 54.8-96.7% increase in the total phenolic content, 38.1-76.2% increase in the total flavonoid content, and 56.7-96.7% increase in the total reducing power when compared with the nontransgenic roots ( p < 0.01). DPPH results also revealed that the transgenic hairy roots exhibited a 31.6-50% increase in antioxidant potential, when compared to normal roots. This study addressed the enhancement of secondary metabolite biosynthesis by hairy root induction in L. serriola .

[Induction and in vitro culture of hairy roots of Dianthus caryophyllus and its plant regeneration].

PubMed

Shi, Heping; Zhu, Yuanfeng; Wang, Bei; Sun, Jiangbing; Huang, Shengqin

2014-11-01

To use Agrobacterium rhizogenes-induced hairy roots to create new germplasm of Dianthus caryophyllus, we transformed D. caryophyllus with A. rhizogenes by leaf disc for plant regeneration from hairy roots. The white hairy roots could be induced from the basal surface of leaf explants of D. caryophyllus 12 days after inoculation with A. rhizogenes ATCC15834. The percentage of the rooting leaf explants was about 90% 21 days after inoculation. The hairy roots could grow rapidly and autonomously in liquid or solid phytohormone-free MS medium. The transformation was confirmed by PCR amplification of rol gene of Ri plasmid and silica gel thin-layer chromatography of opines from D. caryophyllus hairy roots. Hairy roots could form light green callus after cultured on MS+6-BA 1.0-3.0 mg/L + NAA 0.1-0.2 mg/L for 15 days. The optimum medium for adventitious shoots formation was MS + 6-BA 2.0 mg/L + NAA 0.02 mg/L, where the rate of adventitious shoot induction was 100% after cultured for 6 weeks. The mean number of adventitious shoot per callus was 30-40. The adventitious shoots can form roots when cultured on phytohormone-free 1/2 MS or 1/2 MS +0.5 mg/L NAA for 10 days. When the rooted plantlets transplanted in the substrate mixed with perlite sand and peat (volume ratio of 1:2), the survival rate was above 95%.

Hairy strings

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sahakian, Vatche

Zero modes of the world-sheet spinors of a closed string can source higher order moments of the bulk supergravity fields. In this work, we analyze various configurations of closed strings focusing on the imprints of the quantized spinor vacuum expectation values onto the tails of bulk fields. We identify supersymmetric arrangements for which all multipole charges vanish; while for others, we find that one is left with Neveu-Schwarz-Neveu-Schwarz, and Ramond-Ramond dipole and quadrupole moments. Our analysis is exhaustive with respect to all the bosonic fields of the bulk and to all higher order moments. We comment on the relevance ofmore » these results to entropy computations of hairy black holes of a single charge or more, and to open/closed string duality.« less

Vortex shedding noise of a cylinder with hairy flaps

NASA Astrophysics Data System (ADS)

Kamps, Laura; Geyer, Thomas F.; Sarradj, Ennes; Brücker, Christoph

2017-02-01

This study describes the modification of acoustic noise emitted from cylinders in a stationary subsonic flow for a cylinder equipped with flexible hairy flaps at the aft part as a passive way to manipulate the flow and acoustics. The study was motivated by the results from previous water tunnel measurements, which demonstrated that hairy flaps can modify the shedding cycle behind the cylinder and can reduce the wake deficit. In the present study, wind tunnel experiments were conducted on such a modified cylinder and the results were compared to the reference case of a plain cylinder. The acoustic spectrum was measured using two microphones while simultaneously recording the flap motion. To further examine the flow structures in the downstream vicinity of the cylinder, constant temperature anemometry measurements as well as flow visualizations were also performed. The results show that, above a certain Reynolds number, the hairy flaps lead to a jump in the vortex shedding frequency. This phenomenon is similarly observed in the water flow experiments as a jump in the non-dimensional Strouhal number that is related to the change of the shedding cycle. This jump appears to be coupled to a resonant excitation of the flaps. The specific Reynolds number at which the jump occurs is higher in the present case, which is attributed to the lower added mass in air as compared with the one in water. The flow visualizations confirmed that such action of the flaps lead to a more slender elongated shape of the time-averaged separation bubble. In addition, the hairy flaps induce a noticeable reduction of the tonal noise as well as broadband noise as long as the flaps do not touch each other.

Hypophosphatemic rickets associated with epidermal nevus syndrome and giant hairy nevus.

PubMed

Chou, Yen-Yin; Chao, Sheau-Chiou; Shiue, Chiou-Nan; Tsai, Wen-Hui; Lin, Shio-Jean

2005-01-01

The association of hypophosphatemic rickets and epidermal nevus or giant hairy nevus is rare. We report two patients with hypophosphatemic rickets, one associated with epidermal nevus syndrome and the other with giant hairy nevus, and describe their clinical features and variable response to treatment. The abnormal nevus tissue may have contributed to the pathogenesis of hypophosphatemic rickets. We did not find a PHEX gene mutation in these two patients, and the mechanism for their rickets may be different from that in X-linked hypophosphatemic rickets.

Genetic Transformation and Hairy Root Induction Enhance the Antioxidant Potential of Lactuca serriola L.

PubMed Central

Elkelish, Amr; Elansary, Hosam O.; Ali, Hayssam M.; Elshikh, Mohamed; Witczak, Jacques; Ahmad, Margaret

2017-01-01

Lactuca serriola L. is a herbaceous species, used for human nutrition and medicinal purposes. The high antioxidant capacity of L. serriola indicates the possibility of enhancing its edible and health potential by increasing the flavonoid and phenolic contents. The present study aimed at enhancing the production of phenolics and flavonoids by hairy root cultures in Lactuca serriola transformed with Agrobacterium rhizogenes strain AR15834 harbouring the rolB gene. The genetic transformation of rolB in transformed roots was validated, and rolB expression level was evaluated using real-time qPCR analysis. Expression levels of flavonoid biosynthesis genes (CHI, PAL, FLS, and CHS) were assessed in the hairy and nontransformed roots. Results showed higher expression levels in the transgenic roots than in the nontransformed ones (p < 0.01). Transgenic hairy roots exhibited a 54.8–96.7% increase in the total phenolic content, 38.1–76.2% increase in the total flavonoid content, and 56.7–96.7% increase in the total reducing power when compared with the nontransgenic roots (p < 0.01). DPPH results also revealed that the transgenic hairy roots exhibited a 31.6–50% increase in antioxidant potential, when compared to normal roots. This study addressed the enhancement of secondary metabolite biosynthesis by hairy root induction in L. serriola. PMID:28835782

Plant hairy root cultures as plasmodium modulators of the slime mold emergent computing substrate Physarum polycephalum.

PubMed

Ricigliano, Vincent; Chitaman, Javed; Tong, Jingjing; Adamatzky, Andrew; Howarth, Dianella G

2015-01-01

Roots of the medicinal plant Valeriana officinalis are well-studied for their various biological activities. We applied genetically transformed V. officinalis root biomass to exert control of Physarum polycephalum, an amoeba-based emergent computing substrate. The plasmodial stage of the P. polycephalum life cycle constitutes a single, multinucleate cell visible by unaided eye. The plasmodium modifies its network of oscillating protoplasm in response to spatial configurations of attractants and repellents, a behavior that is interpreted as biological computation. To program the computing behavior of P. polycephalum, a diverse and sustainable library of plasmodium modulators is required. Hairy roots produced by genetic transformation with Agrobacterium rhizogenes are a metabolically stable source of bioactive compounds. Adventitious roots were induced on in vitro V. officinalis plants following infection with A. rhizogenes. A single hairy root clone was selected for massive propagation and the biomass was characterized in P. polycephalum chemotaxis, maze-solving, and electrical activity assays. The Agrobacterium-derived roots of V. officinalis elicited a positive chemotactic response and augmented maze-solving behavior. In a simple plasmodium circuit, introduction of hairy root biomass stimulated the oscillation patterns of slime mold's surface electrical activity. We propose that manipulation of P. polycephalum with the plant root culture platform can be applied to the development of slime mold microfluidic devices as well as future models for engineering the plant rhizosphere.

Plant hairy root cultures as plasmodium modulators of the slime mold emergent computing substrate Physarum polycephalum

PubMed Central

Ricigliano, Vincent; Chitaman, Javed; Tong, Jingjing; Adamatzky, Andrew; Howarth, Dianella G.

2015-01-01

Roots of the medicinal plant Valeriana officinalis are well-studied for their various biological activities. We applied genetically transformed V. officinalis root biomass to exert control of Physarum polycephalum, an amoeba-based emergent computing substrate. The plasmodial stage of the P. polycephalum life cycle constitutes a single, multinucleate cell visible by unaided eye. The plasmodium modifies its network of oscillating protoplasm in response to spatial configurations of attractants and repellents, a behavior that is interpreted as biological computation. To program the computing behavior of P. polycephalum, a diverse and sustainable library of plasmodium modulators is required. Hairy roots produced by genetic transformation with Agrobacterium rhizogenes are a metabolically stable source of bioactive compounds. Adventitious roots were induced on in vitro V. officinalis plants following infection with A. rhizogenes. A single hairy root clone was selected for massive propagation and the biomass was characterized in P. polycephalum chemotaxis, maze-solving, and electrical activity assays. The Agrobacterium-derived roots of V. officinalis elicited a positive chemotactic response and augmented maze-solving behavior. In a simple plasmodium circuit, introduction of hairy root biomass stimulated the oscillation patterns of slime mold's surface electrical activity. We propose that manipulation of P. polycephalum with the plant root culture platform can be applied to the development of slime mold microfluidic devices as well as future models for engineering the plant rhizosphere. PMID:26236301

[Role of NO signal in ABA-induced phenolic acids accumulation in Salvia miltiorrhiza hairy roots].

PubMed

Shen, Lihong; Ren, Jiahui; Jin, Wenfang; Wang, Ruijie; Ni, Chunhong; Tong, Mengjiao; Liang, Zongsuo; Yang, Dongfeng

2016-02-01

To investigate roles of nitric oxide (NO) signal in accumulations of phenolic acids in abscisic.acid (ABA)-induced Salvia miltiorrhiza hairy roots, S. miltiorrhiza hairy roots were treated with different concentrations of sodium nitroprusside (SNP)-an exogenous NO donor, for 6 days, and contents of phenolic acids in the hairy roots are determined. Then with treatment of ABA and NO scavenger (2-(4-carboxy-2-phenyl)-4,4,5,5-tetramethylimidazoline-1- oxyl-3-oxide, c-PTIO) or NO synthase inhibitor (NG-nitro-L-arginine methyl ester, L-NAME), contents of phenolic acids and expression levels of three key genes involved in phenolic acids biosynthesis were detected. Phenolic acids production in S. miltiorrhiza hairy roots was most significantly improved by 100 µmoL/L SNP. Contents of RA and salvianolic acid B increased by 3 and 4 folds. ABA significantly improved transcript levels of PAL (phenylalanine ammonia lyase), TAT (tyrosine aminotransferase) and RAS (rosmarinic acid synthase), and increased phenolic acids accumulations. However, with treatments of ABA+c-PTIO or ABA+L-NAME, accumulations of phenolic acids and expression levels of the three key genes were significantly inhibited. Both NO and ABA can increase accumulations of phenolic acids in S. miltiorrhiza hairy roots. NO signal probably mediates the ABA-induced phenolic acids production.

Validation of a hairy roots system to study soybean-soybean aphid interactions

PubMed Central

Morriss, Stephanie C.; Studham, Matthew E.; Tylka, Gregory L.

2017-01-01

The soybean aphid (Aphis glycines) is one of the main insect pests of soybean (Glycine max) worldwide. Genomics approaches have provided important data on transcriptome changes, both in the insect and in the plant, in response to the plant-aphid interaction. However, the difficulties to transform soybean and to rear soybean aphid on artificial media have hindered our ability to systematically test the function of genes identified by those analyses as mediators of plant resistance to the insect. An efficient approach to produce transgenic soybean material is the production of transformed hairy roots using Agrobacterium rhizogenes; however, soybean aphids colonize leaves or stems and thus this approach has not been utilized. Here, we developed a hairy root system that allowed effective aphid feeding. We show that this system supports aphid performance similar to that observed in leaves. The use of hairy roots to study plant resistance is validated by experiments showing that roots generated from cotyledons of resistant lines carrying the Rag1 or Rag2 resistance genes are also resistant to aphid feeding, while related susceptible lines are not. Our results demonstrate that hairy roots are a good system to study soybean aphid-soybean interactions, providing a quick and effective method that could be used for functional analysis of the resistance response to this insect. PMID:28358854

Elicitation Based Enhancement of Secondary Metabolites in Rauwolfia serpentina and Solanum khasianum Hairy Root Cultures.

PubMed

Srivastava, Mrinalini; Sharma, Swati; Misra, Pratibha

2016-05-01

Rauwolfia serpentina and Solanum khasianum are well-known medicinally important plants contained important alkaloids in their different parts. Elicitation of these alkaloids is important because of associated pharmaceutical properties. Targeted metabolites were ajmaline and ajmalicine in R. serpentina; solasodine and α-solanine in S. khasianum. Enhancement of secondary metabolites through biotic and abiotic elicitors in hairy root cultures of R. serpentina and S. khasianum. In this report, hairy root cultures of these two plants were established through Agrobacterium rhizogenes mediated transformation by optimizing various parameters as age of explants, duration of preculture, and co-cultivation period. NaCl was used as abiotic elicitors in these two plants. Cellulase from Aspergillus niger was used as biotic elicitor in S. khasianum and mannan from Saccharomyces cerevisiae was used in R. serpentina. First time we have reported the effect of biotic and abiotic elicitors on the production of important metabolites in hairy root cultures of these two plants. Ajmalicine production was stimulated up to 14.8-fold at 100 mM concentration of NaCl after 1 week of treatment. Ajmaline concentration was also increased 2.9-fold at 100 mg/l dose of mannan after 1 week. Solasodine content was enhanced up to 4.0-fold and 3.6-fold at 100 mM and 200 mM NaCl, respectively, after 6 days of treatments. This study explored the potential of the elicitation strategy in A. rhizogenes transformed cell cultures and this potential further used for commercial production of these pharmaceutically important secondary metabolites. Hairy roots of Rauwolfia serpentina were subjected to salt (abiotic stress) and mannan (biotic stress) treatment for 1 week. Ajmaline and ajmalicine secondary metabolites were quantified before and after stress treatmentAjmalicine yield was enhanced up to 14.8-fold at 100 mM concentration of NaCl. Ajmaline content was also stimulated 2.9-fold at 100 mg/l dose of mannan

Metabolic shift from withasteroid formation to phenylpropanoid accumulation in cryptogein-cotransformed hairy roots of Withania somnifera (L.) Dunal.

PubMed

Sil, Bipradut; Mukherjee, Chiranjit; Jha, Sumita; Mitra, Adinpunya

2015-07-01

Cotransformed hairy roots containing a gene that encodes a fungal elicitor protein, β-cryptogein, were established in Withania somnifera, a medicinal plant widely used in Indian systems of medicine. To find out whether β-cryptogein protein endogenously elicits the pathway of withasteroid biosynthesis, withaferin A and withanolide A contents along with transcript accumulation of farnesyl pyrophosphate (FPP) synthase, 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGR), and sterol glycosyltransferase (SGT) were analyzed in both cryptogein-cotransformed and normal hairy roots of W. somnifera. It was observed that the withaferin A and withanolide A contents were drastically higher in normal hairy roots than cryptogein-cotransformed ones. Similar trends were also observed on the levels of transcript accumulation. Subsequently, the enzyme activity of phenylalanine ammonia lyase (PAL), one of the key enzymes of phenylpropanoid pathway, was measured in both cryptogein-cotransformed and normal hairy roots of W. somnifera along with the levels of PAL transcript accumulation. Upliftment of PAL activity was observed in cryptogein-cotransformed hairy roots as compared to the normal ones, and the PAL expression also reflected a similar trend, i.e., enhanced expression in the cryptogein-cotransformed lines. Upliftment of wall-bound ferulic acid accumulation was also observed in the cryptogein-cotransformed lines, as compared to normal hairy root lines. Thus, the outcome of the above studies suggests a metabolic shift from withanolide accumulation to phenylpropanoid biosynthesis in cryptogein-cotransformed hairy roots of W. somnifera.

High-efficiency induction of soybean hairy roots and propagation of the soybean cyst nematode.

PubMed

Cho, H J; Farrand, S K; Noel, G R; Widholm, J M

2000-01-01

Cotyledon explants of 10 soybean [Glycine max (L.) Merr.] cultivars were inoculated with Agrobacterium rhizogenes strain K599 with and without binary vectors pBI121 or pBINm-gfp5-ER possessing both neomycin phosphotransferase II (nptII) and beta-glucuronidase (gus) or nptII and green fluorescent protein (gfp) genes, respectively. Hairy roots were produced from the wounded surface of 54-95% of the cotyledon explants on MXB selective medium containing 200 microg ml(-1) kanamycin and 500 microg ml(-1) carbenicillin. Putative individual transformed hairy roots were identified by cucumopine analysis and were screened for transgene incorporation using polymerase chain reaction. All of the roots tested were found to be co-transformed with T-DNA from the Ri-plasmid and the transgene from the binary vectors. Southern blot analysis confirmed the presence of the 35S-gfp5 gene in the plant genomes. Transgene expression was also confirmed by histochemical GUS assay and Western blot analysis for the GFP. Attempts to induce shoot formation from the hairy roots failed. Infection of hairy roots of the soybean cyst nematode (Heterodera glycines Ichinohe)-susceptible cultivar, Williams 82, with eggs of H. glycines race 1, resulted in the development of mature cysts about 4-5 weeks after inoculation. Thus the soybean cyst nematode could complete its entire life cycle in transformed soybean hairy-root cultures expressing GFP. This system should be ideal for testing genes that might impart resistance to soybean cyst nematode.

Optimal inductive and cultural conditions of Polygonum multiflorum transgenic hairy roots mediated with Agrobacterium rhizogenes R1601 and an analysis of their anthraquinone constituents.

PubMed

Huang, Bing; Lin, Huanjie; Yan, Chuanyan; Qiu, Hongyan; Qiu, Lipeng; Yu, Rongmin

2014-01-01

Polygonum multiflorum is an important medicinal plant. Hairy roots systems obtained by transforming plant tissues with the natural genetic engineer Agrobacterium rhizogenes can produce valuable biological active substances, which have immense potential in the pharmaceutical industry. To optimize the inductive and cultural conditions of P. multiflorum hairy roots and to identify the major active secondary metabolites in hairy roots. P. multiflorum hairy root were mediated with A. rhizogenes R1601 to induce hairy roots. Four combinations, including Murashige-Skoog (MS), 1/2 MS, B5, and White, were investigated to optimize the culture medium. MS medium was selected for the growth measurement. The qualitative and quantitative determinations of free anthraquinone in hairy roots were compared with the calli and aseptic plantlets using high-performance liquid chromatography. The inductive rates of hairy roots by leaves were higher than for any other explants. The presence of agropine in the P. multiflorum hairy roots confirmed that they were indeed transgenic. MS medium was the most suitable of the four media for hairy root growth. Meanwhile, the growth kinetics and nutrient consumption results showed that the hairy roots displayed a sigmoidal growth curve and that their optimal inoculation time was 18-21 days. The determination of the anthraquinone constituents indicated that the rhein content of the hairy roots reached 2.495 μg g(-1) and was 2.55-fold higher than that of natural plants. Transgenic hairy roots of P. multiflorum could be one of the most potent materials for industrial-scale production of bioactive anthraquinone constituents.

Hairy root biotechnology of Rauwolfia serpentina: a potent approach for the production of pharmaceutically important terpenoid indole alkaloids.

PubMed

Mehrotra, Shakti; Goel, Manoj K; Srivastava, Vikas; Rahman, Laiq Ur

2015-02-01

Hairy root cultures of Rauwolfia serpentina induced by Agrobacterium rhizogenes have been investigated extensively for the production of terpenoid indole alkaloids. Various biotechnological developments, such as scaling up in bioreactors, pathway engineering etc., have been explored to improve their metabolite production potential. These hairy roots are competent for regenerating into complete plants and show survival and unaltered biosynthetic potential during storage at low temperature. This review provides a comprehensive account of the hairy root cultures of R. serpentina, their biosynthetic potential and various biotechnological methods used to explore the production of pharmaceutically important terpenoid indole alkaloids. The review also indicates how biotechnological endeavors might improve the future progress of research for production of alkaloids using Rauwolfia hairy roots.

Precisely Size-Tunable Monodisperse Hairy Plasmonic Nanoparticles via Amphiphilic Star-Like Block Copolymers.

PubMed

Chen, Yihuang; Yoon, Young Jun; Pang, Xinchang; He, Yanjie; Jung, Jaehan; Feng, Chaowei; Zhang, Guangzhao; Lin, Zhiqun

2016-12-01

In situ precision synthesis of monodisperse hairy plasmonic nanoparticles with tailored dimensions and compositions by capitalizing on amphiphilic star-like diblock copolymers as nanoreactors are reported. Such hairy plasmonic nanoparticles comprise uniform noble metal nanoparticles intimately and perpetually capped by hydrophobic polymer chains (i.e., "hairs") with even length. Interestingly, amphiphilic star-like diblock copolymer nanoreactors retain the spherical shape under reaction conditions, and the diameter of the resulting plasmonic nanoparticles and the thickness of polymer chains situated on the surface of the nanoparticle can be readily and precisely tailored. These hairy nanoparticles can be regarded as hard/soft core/shell nanoparticles. Notably, the polymer "hairs" are directly and permanently tethered to the noble metal nanoparticle surface, thereby preventing the aggregation of nanoparticles and rendering their dissolution in nonpolar solvents and the homogeneous distribution in polymer matrices with long-term stability. This amphiphilic star-like block copolymer nanoreactor-based strategy is viable and robust and conceptually enables the design and synthesis of a rich variety of hairy functional nanoparticles with new horizons for fundamental research on self-assembly and technological applications in plasmonics, catalysis, energy conversion and storage, bioimaging, and biosensors. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Establishment of Withania somnifera hairy root cultures for the production of withanolide A.

PubMed

Murthy, Hosakatte N; Dijkstra, Camelia; Anthony, Paul; White, Daniel A; Davey, Mike R; Power, J Brian; Hahn, Eun J; Paek, Kee Y

2008-08-01

Withania sominifera (Indian ginseng) was transformed by Agrobacterium rhizogenes. Explants from seedling roots, stems, hypocotyls, cotyledonary nodal segments, cotyledons and young leaves were inoculated with A. rhizogenes strain R1601. Hairy (transformed) roots were induced from cotyledons and leaf explants. The transgenic status of hairy roots was confirmed by polymerase chain reaction using nptII and rolB specific primers and, subsequently, by Southern analysis for the presence of nptII and rolB genes in the genomes of transformed roots. Four clones of hairy roots were established; these differed in their morphology. The doubling time of faster growing cultures was 8-14 d with a fivefold increase in biomass after 28 d compared with cultured, non-transformed seedling roots. MS-based liquid medium was superior for the growth of transformed roots compared with other culture media evaluated (SH, LS and N6), with MS-based medium supplemented with 40 g/L sucrose being optimal for biomass production. Cultured hairy roots synthesized withanolide A, a steroidal lactone of medicinal and therapeutic value. The concentration of withanolide A in transformed roots (157.4 microg/g dry weight) was 2.7-fold more than in non-transformed cultured roots (57.9 microg/g dry weight).

In vitro azadirachtin production by hairy root cultivation of Azadirachta indica in nutrient mist bioreactor.

PubMed

Srivastava, Smita; Srivastava, A K

2012-01-01

Azadirachtin, a well-known biopesticide is a secondary metabolite conventionally extracted from the seeds of Azadirachta indica. The present study involved in vitro azadirachtin production by developing hairy roots of A. indica via Agrobacterium rhizogenes-mediated transformation of A. indica explants. Liquid culture of hairy roots was established in shake flask to study the kinetics of growth and azadirachtin production. A biomass production of 13.3 g/L dry weight (specific growth rate of 0.7 day(-1)) was obtained after 25 days of cultivation period with an azadirachtin yield of 3.3 mg/g root biomass. To overcome the mass transfer limitation in conventionally used liquid-phase reactors, batch cultivation of hairy roots was carried out in gas-phase reactors (nutrient spray and nutrient mist bioreactor) to investigate the possible scale-up of A. indica hairy root culture. The nano-size nutrient mist particles generated from the nozzle of the nutrient mist bioreactor could penetrate till the inner core of the inoculated root matrix, facilitating uniform growth during high-density cultivation of hairy roots. A biomass production of 9.8 g/L dry weight with azadirachtin accumulation of 2.8 mg/g biomass (27.4 mg/L) could be achieved in 25 days of batch cultivation period, which was equivalent to a volumetric productivity of 1.09 mg/L per day of azadirachtin.

Overexpression of cinnamate 4-hydroxylase gene enhances biosynthesis of decursinol angelate in Angelica gigas hairy roots.

PubMed

Park, Nam Il; Park, Jee Hee; Park, Sang Un

2012-02-01

Angelica gigas is a medicinal plant that produces pyranocoumarins, including decursin (D) and decursinol angelate (DA), which have neuroprotective, anticancer, and antiandrogenic effects. In this study, the coumarin biosynthetic pathway was engineered to increase the production of DA. Specifically, a vector was constructed which contained the A. gigas phenylalanine ammonia-lyase (AgPAL) and cinnamate 4-hydroxylase (AgC4H) genes that were driven by the cauliflower mosaic virus (CaMV) 35S promoter. Transgenic hairy roots that overexpressed AgPAL or AgC4H genes were obtained by using an Agrobacterium rhizogenes-mediated transformation system. Among them, only AgC4H-transgenic hairy root lines produced more DA than control transgenic hairy root lines. The enhanced gene expression corresponded to elevated C4H activities. This study showed the importance of C4H in the production of DA in A. gigas hairy root culture.

Use of Chenopodium murale L. transgenic hairy root in vitro culture system as a new tool for allelopathic assays.

PubMed

Mitić, Nevena; Dmitrović, Slavica; Djordjević, Mirka; Zdravković-Korać, Snežana; Nikolić, Radomirka; Raspor, Martin; Djordjević, Tatjana; Maksimović, Vuk; Zivković, Suzana; Krstić-Milošević, Dijana; Stanišić, Mariana; Ninković, Slavica

2012-08-15

We investigated Chenopodium murale transgenic hairy root in vitro culture system as a new tool for allelopathic assays. Transgenic hairy roots were induced by Agrobacterium rhizogenes A4M70GUS from roots, cotyledons, leaves, and internodes of C. murale seedlings. Roots were found to be the best target explants, providing transformation efficiency of up to 11.1%. Established hairy root clones differed in their morphology and growth potential. Molecular characterization of these clones was carried out by PCR, RT-PCR and histochemical GUS analyses. No differences in rol gene expression were observed. Liquid culture system of characterized hairy root clones was maintained for over 2 years. Six hairy root clones were selected for assaying the allelopathic effect of their growth medium against germination and seedling elongation of wheat and lettuce test plants. The inhibitory potential varied depending on the hairy root clone. Some transgenic clones showed significantly higher inhibition compared to wild-type roots. These results revealed that hairy roots as an independent system synthesize some bioactive substances with allelopathic activity and exude them into the growth medium. Concentrations of caffeic, ferulic and p-coumaric acids (0.07-2.85 μmol/L) identified by HPLC analysis in the growth media were at least 1000 times lower than the inhibitory active concentration (5 mmol/L) of pure grade phenolic acids, suggesting that they have a limited role in the allelopathic phenomena of C. murale. The presented hairy root system appears to be a suitable tool for further investigation of the potential and nature of root-mediated allelopathic interference of C. murale. Copyright © 2012 Elsevier GmbH. All rights reserved.

Small RNAs Derived from the T-DNA of Agrobacterium rhizogenes in Hairy Roots of Phaseolus vulgaris

PubMed Central

Peláez, Pablo; Hernández-López, Alejandrina; Estrada-Navarrete, Georgina; Sanchez, Federico

2017-01-01

Agrobacterium rhizogenes is a pathogenic bacteria that causes hairy root disease by transferring bacterial DNA into the plant genome. It is an essential tool for industry and research due to its capacity to produce genetically modified roots and whole organisms. Here, we identified and characterized small RNAs generated from the transfer DNA (T-DNA) of A. rhizogenes in hairy roots of common bean (Phaseolus vulgaris). Distinct abundant A. rhizogenes T-DNA-derived small RNAs (ArT-sRNAs) belonging to several oncogenes were detected in hairy roots using high-throughput sequencing. The most abundant and diverse species of ArT-sRNAs were those of 21- and 22-nucleotides in length. Many T-DNA encoded genes constituted phasiRNA producing loci (PHAS loci). Interestingly, degradome analysis revealed that ArT-sRNAs potentially target genes of P. vulgaris. In addition, we detected low levels of ArT-sRNAs in the A. rhizogenes-induced calli generated at the wound site before hairy root emergence. These results suggest that RNA silencing targets several genes from T-DNA of A. rhizogenes in hairy roots of common bean. Therefore, the role of RNA silencing observed in this study has implications in our understanding and usage of this unique plant-bacteria interaction. PMID:28203245

A novel life cycle arising from leaf segments in plants regenerated from horseradish hairy roots.

PubMed

Mano, Y; Matsuhashi, M

1995-03-01

Horseradish (Armoracia rusticana) hairy root clones were established from hairy roots which were transformed with the Ri plasmid in Agrobacterium rhizogenes 15834. The transformed plants, which were regenerated from hairy root clones, had thicker roots with extensive lateral branches and thicker stems, and grew faster compared with non-transformed horseradish plants. Small sections of leaves of the transformed plants generated adventitious roots in phytohormone-free G (modified Gamborg's) medium. Root proliferation was followed by adventitious shoot formation and plant regeneration. Approximately twenty plants were regenerated per square centimeter of leaf. The transformed plants were easily transferable from sterile conditions to soil. When leaf segments of the transformed plants were cultured in a liquid fertilizer under non-sterile conditions, adventitious roots were generated at the cut ends of the leaves. Adventitious shoots were generated at the boundary between the leaf and the adventitious roots and developed into complete plants. This novel life cycle arising from leaf segments is a unique property of the transformed plants derived from hairy root clones.

Induction of hairy roots by various strains of Agrobacterium rhizogenes in different types of Capsicum species explants.

PubMed

Md Setamam, Nursuria; Jaafar Sidik, Norrizah; Abdul Rahman, Zainon; Che Mohd Zain, Che Radziah

2014-06-30

Capsicum annuum and Capsicum frutescens, also known as "chilies", belong to the Solanaceae family and have tremendous beneficial properties. The application of hairy root culture may become an alternative method for future development of these species by adding value, such as by increasing secondary metabolites and improving genetic and biochemical stability compared with normal Capsicum plants. Therefore, in this research, different types of explants of both species were infected with various Agrobacterium rhizogenes strains to provide more information about the morphology and induction efficiency of hairy roots. After 2 weeks of in vitro seed germination, young seedling explants were cut into three segments; the cotyledon, hypocotyl, and radical. Then, the explants were co-cultured with four isolated A. rhizogenes strains in Murashige & Skoog culture media (MS) containing decreasing carbenicillin disodium concentrations for one month. In this experiment, thick and short hairy roots were induced at all induction sites of C. annuum while thin, elongated hairy roots appeared mostly at wound sites of C. frutescens. Overall, the hairy root induction percentages of C. frutescens were higher compared with C. annuum. Hairy root initiation was observed earliest using radicles (1st week), followed by cotyledons (2nd week), and hypocotyls (3rd week). Cotyledon explants of both species had the highest induction frequency with all strains compared with the other explants types. Strains ATCC 13333 and ATCC 15834 were the most favourable for C. frutescens while ATCC 43056 and ATCC 43057 were the most favourable for C. annuum. The interactions between the different explants and strains showed significant differences with p-values < 0.0001 in both Capsicum species. Both Capsicum species were amenable to A. rhizogenes infection and hairy root induction is recommended for use as an alternative explants in future plant-based studies.

Induction of hairy roots by various strains of Agrobacterium rhizogenes in different types of Capsicum species explants

PubMed Central

2014-01-01

Background Capsicum annuum and Capsicum frutescens, also known as “chilies”, belong to the Solanaceae family and have tremendous beneficial properties. The application of hairy root culture may become an alternative method for future development of these species by adding value, such as by increasing secondary metabolites and improving genetic and biochemical stability compared with normal Capsicum plants. Therefore, in this research, different types of explants of both species were infected with various Agrobacterium rhizogenes strains to provide more information about the morphology and induction efficiency of hairy roots. After 2 weeks of in vitro seed germination, young seedling explants were cut into three segments; the cotyledon, hypocotyl, and radical. Then, the explants were co-cultured with four isolated A. rhizogenes strains in Murashige & Skoog culture media (MS) containing decreasing carbenicillin disodium concentrations for one month. Results In this experiment, thick and short hairy roots were induced at all induction sites of C. annuum while thin, elongated hairy roots appeared mostly at wound sites of C. frutescens. Overall, the hairy root induction percentages of C. frutescens were higher compared with C. annuum. Hairy root initiation was observed earliest using radicles (1st week), followed by cotyledons (2nd week), and hypocotyls (3rd week). Cotyledon explants of both species had the highest induction frequency with all strains compared with the other explants types. Strains ATCC 13333 and ATCC 15834 were the most favourable for C. frutescens while ATCC 43056 and ATCC 43057 were the most favourable for C. annuum. The interactions between the different explants and strains showed significant differences with p-values < 0.0001 in both Capsicum species. Conclusions Both Capsicum species were amenable to A. rhizogenes infection and hairy root induction is recommended for use as an alternative explants in future plant-based studies. PMID

The shikonin derivatives and pyrrolizidine alkaloids in hairy root cultures of Lithospermum canescens (Michx.) Lehm.

PubMed

Pietrosiuk, A; Sykłowska-Baranek, K; Wiedenfeld, H; Wolinowska, R; Furmanowa, M; Jaroszyk, E

2006-10-01

Hairy root cultures of Lithospermum canescens were established using three strains of Agrobacterium rhizogenes: ATCC 15834, LBA 9402 and NCIB 8196. Eight lines resulting from infection with A. rhizogenes ATCC 15834 demonstrated sufficient biomass increase and were submitted to further investigations. The contents of acetylshikonin (ACS) and isobutyrylshikonin (IBS) in transformed hairy roots made up ca. 10% of those observed in natural roots of L. canescens (24.35 and 14.48 mg g(-1) DW, respectively). One line, Lc1-D, produced the largest amounts of ACS (2.72 mg g(-1) DW) and IBS (0.307 mg g(-1) DW). Traces of pyrrolizidine alkaloids (PA), canescine and canescenine, were found in all lines of transformed hairy roots.

Artemisinin production by plant hairy root cultures in gas- and liquid-phase bioreactors.

PubMed

Patra, Nivedita; Srivastava, Ashok K

2016-01-01

Alternative biotechnological protocol for large-scale artemisinin production was established. It featured enhanced growth and artemisinin production by cultivation of hairy roots in nutrient mist bioreactor (NMB) coupled with novel cultivation strategies. Artemisinin is used for the treatment of cerebral malaria. Presently, its main source is from seasonal plant Artemisia annua. This study featured investigation of growth and artemisinin production by A. annua hairy roots (induced by Agrobacterium rhizogenes-mediated genetic transformation of explants) in three bioreactor configurations-bubble column reactor, NMB and modified NMB particularly to establish their suitability for commercial production. It was observed that cultivation of hairy roots in a non-stirred bubble column reactor exhibited a biomass accumulation of 5.68 g/l only while batch cultivation in a custom-made NMB exhibited a higher biomass concentration of 8.52 g/l but relatively lower artemisinin accumulation of 0.22 mg/g was observed in this reactor. A mixture of submerged liquid-phase growth (for 5 days) followed by gas-phase cultivation in nutrient mist reactor operation strategy (for next 15 days) was adopted for hairy root cultivation in this investigation. Reasonably, high (23.02 g/l) final dry weight along with the artemisinin accumulation (1.12 mg/g, equivalent to 25.78 mg/l artemisinin) was obtained in this bioreactor, which is the highest reported artemisinin yield in the gas-phase NMB cultivation.

Habitat Suitability Index Models: Hairy woodpecker

USGS Publications Warehouse

Sousa, Patrick J.

1987-01-01

A review and synthesis of existing information were used to develop a Habitat Suitability Index (HSI) model for the hairy woodpecker (Picoides villosus). The model consolidates habitat use information into a framework appropriate for field application, and is scaled to produce an index between 0.0 (unsuitable habitat) to 1.0 (optimum habitat). HSI models are designed to be used with Habitat Evaluation Procedures previously developed by the U.S. Fish and Wildlife Service.

Hairy black holes and duality in an extended supergravity model

NASA Astrophysics Data System (ADS)

Anabalón, Andrés; Astefanesei, Dumitru; Gallerati, Antonio; Trigiante, Mario

2018-04-01

We consider a D = 4, N=2 gauged supergravity with an electromagnetic Fayet-Iliopoulos term. We restrict to the uncharged, single dilaton consistent truncation and point out that the bulk Lagrangian is self-dual under electromagnetic duality. Within this truncation, we construct two families of exact hairy black hole solutions, which are asymptotically AdS 4. When a duality transformation is applied on these solutions, they are mapped to two other inequivalent families of hairy black hole solutions. The mixed boundary conditions of the scalar field correspond to adding a triple-trace operator to the dual field theory action. We also show that this truncation contains all the consistent single dilaton truncations of gauged N=8 supergravity with a possible ω-deformation.

Elicitation Based Enhancement of Secondary Metabolites in Rauwolfia serpentina and Solanum khasianum Hairy Root Cultures

PubMed Central

Srivastava, Mrinalini; Sharma, Swati; Misra, Pratibha

2016-01-01

Background: Rauwolfia serpentina and Solanum khasianum are well-known medicinally important plants contained important alkaloids in their different parts. Elicitation of these alkaloids is important because of associated pharmaceutical properties. Targeted metabolites were ajmaline and ajmalicine in R. serpentina; solasodine and α-solanine in S. khasianum. Objective: Enhancement of secondary metabolites through biotic and abiotic elicitors in hairy root cultures of R. serpentina and S. khasianum. Materials and Methods: In this report, hairy root cultures of these two plants were established through Agrobacterium rhizogenes mediated transformation by optimizing various parameters as age of explants, duration of preculture, and co-cultivation period. NaCl was used as abiotic elicitors in these two plants. Cellulase from Aspergillus niger was used as biotic elicitor in S. khasianum and mannan from Saccharomyces cerevisiae was used in R. serpentina. Results: First time we have reported the effect of biotic and abiotic elicitors on the production of important metabolites in hairy root cultures of these two plants. Ajmalicine production was stimulated up to 14.8-fold at 100 mM concentration of NaCl after 1 week of treatment. Ajmaline concentration was also increased 2.9-fold at 100 mg/l dose of mannan after 1 week. Solasodine content was enhanced up to 4.0-fold and 3.6-fold at 100 mM and 200 mM NaCl, respectively, after 6 days of treatments. Conclusion: This study explored the potential of the elicitation strategy in A. rhizogenes transformed cell cultures and this potential further used for commercial production of these pharmaceutically important secondary metabolites. SUMMARY Hairy roots of Rauwolfia serpentina were subjected to salt (abiotic stress) and mannan (biotic stress) treatment for 1 week. Ajmaline and ajmalicine secondary metabolites were quantified before and after stress treatmentAjmalicine yield was enhanced up to 14.8-fold at 100 mM concentration of Na

75 FR 14391 - Diseases Associated With Exposure to Certain Herbicide Agents (Hairy Cell Leukemia and Other...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-03-25

... service connection for certain diseases based upon the most recent National Academy of Sciences (NAS... proposed amendment is to establish presumptive service connection for these diseases based on herbicide... Agent Orange Act of 1991, Public Law 102-4, provides that whenever the Secretary determines, based on...

Enhancement of ginsenoside Rg(1) in Panax ginseng hairy root by overexpressing the α-L-rhamnosidase gene from Bifidobacterium breve.

PubMed

Zhang, Ru; Zhang, Bian-Ling; Li, Gu-Cai; Xie, Tao; Hu, Teng; Luo, Zhi-Yong

2015-10-01

To improve the production of ginsenoside Rg1 in Panax ginseng. The α-L-rhamnosidase gene from Bifidobacterium breve (BbRha) was overexpressed into hairy root culture system using Agrobacterium rhizogenes A4. Ginsenoside Rg1 in hairy roots was obtained following transformation via overexpressed gene representing 2.2-fold higher than those of control lines. Several overexpression transgenic hairy root lines were obtained exhibiting markedly increased levels of the corresponding α-L-rhamnosidase enzymatic activity relative to control. Ginsenoside Rg1 levels in the transgenic lines were higher (2.2-fold) than those of control after following 30 days culturing, while ginsenoside Re contents in tested transgenic lines were found to be lower. The transgenic hairy roots harboring α-L-rhamnosidase gene improved the accumulation of ginsenoside Rg1 up to 3.6 mg g(-1) dry weight. BbRha gene selectively enhances the production of ginsenoside Rg1 in P. ginseng hairy roots.

Long-term follow-up and second malignancies in 487 patients with hairy cell leukaemia.

PubMed

Cornet, Edouard; Tomowiak, Cécile; Tanguy-Schmidt, Aline; Lepretre, Stéphane; Dupuis, Jehan; Feugier, Pierre; Devidas, Alain; Mariette, Clara; Leblond, Véronique; Thiéblemont, Catherine; Validire-Charpy, Patricia; Sutton, Laurent; Gyan, Emmanuel; Eisenmann, Jean-Claude; Cony-Makhoul, Pascale; Ysebaert, Loïc; Troussard, Xavier

2014-08-01

A large, multicentre, retrospective survey of patients with hairy cell leukaemia (HCL) was conducted in France to determine the frequency of second malignancies and to analyse the long-term effects of the established purine nucleoside analogues (PNAs), cladribine and pentostatin. The survey retrospectively reviewed the medical history of patients and their immediate family, clinical and biological presentation at the time of HCL diagnosis, treatment choice, response to treatment, time to relapse and cause of death. Data were collected for 487 patients with HCL. Of the patients included in the survey, 18% (88/487) had a familial history of cancers, 8% (41/487) presented with malignancies before HCL diagnosis and 10% (48/487) developed second malignancies after HCL was diagnosed. An excess incidence of second malignancies was observed, with a standardized incidence ratio (SIR) of 1·86 (95% confidence interval (CI): 1·34-2·51), with no significant difference between PNAs. For second haematological malignancies alone, the SIR was markedly increased at 5·32 (95% CI: 2·90-8·92). This study highlights the high frequency of cancers in HCL patients and their family members. The frequency of second malignancies is notably increased, particularly for haematological malignancies. The respective role of pentostatin and cladribine in the development of second malignancies is debatable. © 2014 John Wiley & Sons Ltd.

Drop Impact on Hairy Surfaces

NASA Astrophysics Data System (ADS)

Nasto, Alice; Hosoi, Anette

2017-11-01

Using a combination of experiments and theory, we investigate the effect of a millimeter-scale hairy texture on impact of liquid drops. By varying the speed of the drop at impact and the spacing of the hairs, we observe a variety of behaviors. For dense hairs and low impact velocity, the liquid drop sits on top of the hair, similar to a Cassie-Baxter state. For higher impact velocity, and intermediate to high density of hairs, the drops penetrate through the surface, but the hairs resist their spreading. For low hair density and high impact velocity, the drops impact and splash.

Keeping warm with fur in cold water: entrainment of air in hairy surfaces

NASA Astrophysics Data System (ADS)

Nasto, Alice; Regli, Marianne; Brun, Pierre-Thomas; Clanet, Christophe; Hosoi, Anette

2015-11-01

Instead of relying on a thick layer of body fat for insulation as many aquatic mammals do, fur seals and otters trap air in their dense fur for insulation in cold water. Using a combination of model experiments and theory, we rationalize this mechanism of air trapping underwater for thermoregulation. For the model experiments, hairy surfaces are fabricated using laser cut molds and casting samples with PDMS. Modeling the hairy texture as a network of capillary tubes, the imbibition speed of water into the hairs is obtained through a balance of hydrostatic pressure and viscous stress. In this scenario, the bending of the hairs and capillary forces are negligible. The maximum diving depth that can be achieved before the hairs are wetted to the roots is predicted from a comparison of the diving speed and imbibition speed. The amount of air that is entrained in hairy surfaces is greater than what is expected for classic Landau-Levich-Derjaguin plate plunging. A phase diagram with the parameters from experiments and biological data allows a comparison of the model system and animals.

Investigation of Linum flavum (L.) Hairy Root Cultures for the Production of Anticancer Aryltetralin Lignans.

PubMed

Renouard, Sullivan; Corbin, Cyrielle; Drouet, Samantha; Medvedec, Barbara; Doussot, Joël; Colas, Cyril; Maunit, Benoit; Bhambra, Avninder S; Gontier, Eric; Jullian, Nathalie; Mesnard, François; Boitel, Michèle; Abbasi, Bilal Haider; Arroo, Randolph R J; Lainé, Eric; Hano, Christophe

2018-03-26

Linum flavum hairy root lines were established from hypocotyl pieces using Agrobacterium rhizogenes strains LBA 9402 and ATCC 15834. Both strains were effective for transformation but induction of hairy root phenotype was more stable with strain ATCC 15834. Whereas similar accumulation patterns were observed in podophyllotoxin-related compounds (6-methoxy-podophyllotoxin, podophyllotoxin and deoxypodophyllotoxin), significant quantitative variations were noted between root lines. The influence of culture medium and various treatments (hormone, elicitation and precursor feeding) were evaluated. The highest accumulation was obtained in Gamborg B5 medium. Treatment with methyl jasmonate, and feeding using ferulic acid increased the accumulation of aryltetralin lignans. These results point to the use of hairy root culture lines of Linum flavum as potential sources for these valuable metabolites as an alternative, or as a complement to Podophyllum collected from wild stands.

Efficient Rutin and Quercetin Biosynthesis through Flavonoids-Related Gene Expression in Fagopyrum tataricum Gaertn. Hairy Root Cultures with UV-B Irradiation

PubMed Central

Huang, Xuan; Yao, Jingwen; Zhao, Yangyang; Xie, Dengfeng; Jiang, Xue; Xu, Ziqin

2016-01-01

Transformed hairy roots had been efficiently induced from the seedlings of Fagopyrum tataricum Gaertn. due to the infection of Agrobacterium rhizogenes. Hairy roots were able to display active elongation with high root branching in 1/2 MS medium without growth regulators. The stable introduction of rolB and aux1 genes of A. rhizogenes WT strain 15834 into F. tataricum plants was confirmed by PCR analysis. Besides, the absence of virD gene confirmed hairy root was bacteria-free. After six different media and different sources of concentration were tested, the culturing of TB7 hairy root line in 1/2 MS liquid medium supplemented with 30 g l-1 sucrose for 20 days resulted in a maximal biomass accumulation (13.5 g l-1 fresh weight, 1.78 g l-1 dry weight) and rutin content (0.85 mg g-1). The suspension culture of hairy roots led to a 45-fold biomass increase and a 4.11-fold rutin content increase in comparison with the suspension culture of non-transformed roots. The transformation frequency was enhanced through preculturing for 2 days followed by infection for 20 min. The UV-B stress treatment of hairy roots resulted in a striking increase of rutin and quercetin production. Furthermore, the hairy root lines of TB3, TB7, and TB28 were chosen to study the specific effects of UV-B on flavonoid accumulation and flavonoid biosynthetic gene expression by qRT-PCR. This study has demonstrated that the UV-B radiation was an effective elicitor that dramatically changed in the transcript abundance of ftpAL, FtCHI, FtCHS, FtF3H, and FtFLS-1 in F. tataricum hairy roots. PMID:26870075

Antioxidant activity of selected stilbenoids and their bioproduction in hairy root cultures of muscadine grape (Vitis rotundifolia Michx.).

PubMed

Nopo-Olazabal, Cesar; Hubstenberger, John; Nopo-Olazabal, Luis; Medina-Bolivar, Fabricio

2013-12-04

Stilbenoids are polyphenolic phytoalexins with health-related properties in humans. Muscadine grape ( Vitis rotundifolia ) hairy root cultures were established via Agrobacterium rhizogenes -mediated transformation, and the effects of growth regulators (3-indolebutyric acid and 6-benzylaminopurine) and methyl jasmonate (MeJA) on stilbenoid production were studied. Twenty-one-day-old hairy root cultures were treated with 100 μM MeJA for 24 h, and then the stilbenoids were extracted from the medium and tissue with ethyl acetate and analyzed by HPLC. Resveratrol, piceid, and ε-viniferin were observed preferentially in tissue, whereas piceatannol was observed only in medium. Growth regulators did not affect the yield of stilbenoids, whereas higher levels were found upon treatment with MeJA. Stilbenoids identified in the hairy root cultures were analyzed for their radical scavenging capacity showing piceatannol and ε-viniferin as the strongest antioxidants. Muscadine grape hairy root cultures were demonstrated to be amenable systems to study stilbenoid biosynthesis and a sustainable source of these bioactive compounds.

Allodynia mediated by C-tactile afferents in human hairy skin.

PubMed

Nagi, Saad S; Rubin, Troy K; Chelvanayagam, David K; Macefield, Vaughan G; Mahns, David A

2011-08-15

We recently showed a contribution of low-threshold cutaneous mechanoreceptors to vibration-evoked changes in the perception of muscle pain. Neutral-touch stimulation (vibration) of the hairy skin during underlying muscle pain evoked an overall increase in pain intensity, i.e. allodynia. This effect appeared to be dependent upon cutaneous afferents, as allodynia was abolished by intradermal anaesthesia. However, it remains unclear whether allodynia results from activation of a single class of cutaneous afferents or the convergence of inputs from multiple classes. Intriguingly, no existing human study has examined the contribution of C-tactile (CT) afferents to allodynia. Detailed psychophysical observations were made in 29 healthy subjects (18 males and 11 females). Sustained muscle pain was induced by infusing hypertonic saline (HS: 5%) into tibialis anterior muscle (TA). Sinusoidal vibration (200 Hz–200 μm) was applied to the hairy skin overlying TA. Pain ratings were recorded using a visual analogue scale (VAS). In order to evaluate the role of myelinated and unmyelinated cutaneous afferents in the expression of vibration-evoked allodynia, compression block of the sciatic nerve, and low-dose intradermal anaesthesia (Xylocaine 0.25%) were used, respectively. In addition, the modulation of muscle pain by gentle brushing (1.0 and 3.0 cm s(−1))--known to excite CT fibres--was examined. Brushing stimuli were applied to the hairy skin with all fibres intact and following the blockade of myelinated afferents. During tonic muscle pain (VAS 4–6), vibration evoked a significant and reproducible increase in muscle pain (allodynia) that persisted following compression of myelinated afferents. During compression block, the sense of vibration was abolished, but the vibration-evoked allodynia persisted. In contrast, selective anaesthesia of unmyelinated cutaneous afferents abolished the allodynia, whereas the percept of vibration remained unaffected. Furthermore

Hairy Root Cultures of Gymnema sylvestre R. Br. to Produce Gymnemic Acid.

PubMed

Rajashekar, J; Kumar, Vadlapudi; Veerashree, V; Poornima, D V; Sannabommaji, Torankumar; Gajula, Hari; Giridhara, B

2016-01-01

Gymnema sylvestre R. Br. (Asclepiadaceae) is an endangered species extensively used in the management of diabetes, obesity, and treatment of various diseases. Uncontrolled exploitation to meet the increasing demand and low seed viability hastens the disappearance of the plant from its natural habitat. Hairy root culture provides a suitable alternative for the enhanced production of active principles. The current protocol provides the optimized culture conditions for the establishment of hairy root cultures and elicitation studies and also confirmation of stable integration of A. rhizogenes plasmid T-DNA into host genetic material by PCR and RT-PCR. Furthermore, it also discusses the suitable methods for the extraction procedures, and qualitative and quantitative analysis of gymnemic acid by HPTLC and HPLC.

Efficient production of flavonoids in Fagopyrum tataricum hairy root cultures with yeast polysaccharide elicitation and medium renewal process

PubMed Central

Zhao, Jiang-Lin; Zou, Liang; Zhang, Cai-Qiong; Li, Yuan-Yuan; Peng, Lian-Xin; Xiang, Da-Bing; Zhao, Gang

2014-01-01

Background: Tartary buckwheat (Fagopyrum tataricum), an excellent edible and medicinal crop, has been widely used as a daily diet and traditional medicine for a long time. The major functional components of Fagopyrum tataricum have been demonstrated to be flavonoids (i.e. rutin and quercetin), which had notable andioxidant, antidiabetic, hypocholesterolemic and antitumor activities. Hairy root culture is a convenient and efficient plant tissue culture system for large scale production of bioactive metabolites. Objective: To enhance the functional flavonoids production in hairy root culture of F. tataricum. Materials and Methods: The elicitation treatment in combination with medium renewal strategy was applied for efficient promoting flavonoids production in F. tataricum hairy root cultures. Results: The exogenous yeast polysaccharide (YPS) elicitor notably stimulated the functional metabolites production in F. tataricum hairy root cultures, and the stimulation effect was concentration-dependent. Combination with the YPS elicitation (200 mg/L) and medium renewal process, the maximal flavonoids yield was enhanced to 47.13 mg/L, about 3.2-fold in comparison with the control culture of 14.88 mg/L. Moreover, this research also revealed the accumulation of these bioactive metabolites resulted from the stimulation of the phenylpropanoid pathway by YPS treatment. These results indicated that the F. tataricum hairy root culture could be an effective system for rutin and quercetin production. PMID:25210309

The slow cell death response when screening chemotherapeutic agents.

PubMed

Blois, Joseph; Smith, Adam; Josephson, Lee

2011-09-01

To examine the correlation between cell death and a common surrogate of death used in screening assays, we compared cell death responses to those obtained with the sulforhodamine B (SRB) cell protein-based "cytotoxicity" assay. With the SRB assay, the Hill equation was used to obtain an IC50 and final cell mass, or cell mass present at infinite agent concentrations, with eight adherent cell lines and four agents (32 agent/cell combinations). Cells were treated with high agent concentrations (well above the SRB IC50) and the death response determined as the time-dependent decrease in cells failing to bind both annexin V and vital fluorochromes by flow cytometry. Death kinetics were categorized as fast (5/32) (similar to the reference nonadherent Jurkat line), slow (17/32), or none (10/32), despite positive responses in the SRB assay in all cases. With slow cell death, a single exposure to a chemotherapeutic agent caused a slow, progressive increase in dead (necrotic) and dying (apoptotic) cells for at least 72 h. Cell death (defined by annexin and/or fluorochrome binding) did not correlate with the standard SRB "cytotoxicity" assay. With the slow cell death response, a single exposure to an agent caused a slow conversion from vital to apoptotic and necrotic cells over at least 72 h (the longest time point examined). Here, increasing the time of exposure to agent concentrations modestly above the SRB IC50 provides a method of maximizing cell kill. If tumors respond similarly, sustained low doses of chemotherapeutic agents, rather than a log-kill, maximum tolerated dose strategy may be an optimal strategy of maximizing tumor cell death.

Establishment of Hairy Root Cultures of Rhaponticum carthamoides (Willd.) Iljin for the Production of Biomass and Caffeic Acid Derivatives

PubMed Central

Skała, Ewa; Kicel, Agnieszka; Olszewska, Monika A.; Kiss, Anna K.

2015-01-01

The aim of the study was to obtain transformed roots of Rhaponticum carthamoides and evaluate their phytochemical profile. Hairy roots were induced from leaf explants by the transformation of Agrobacterium rhizogenes strains A4 and ATCC 15834. The best response (43%) was achieved by infection with A4 strain. The effects of different liquid media (WPM, B5, SH) with full and half-strength concentrations of macro- and micronutrients on biomass accumulation of the best grown hairy root line (RC3) at two different lighting conditions (light or dark) were investigated. The highest biomass (93 g L−1 of the fresh weight after 35 days) was obtained in WPM medium under periodic light. UPLC-PDA-ESI-MS3 and HPLC-PDA analyses of 80% aqueous methanol extracts from the obtained hairy roots revealed the presence of eleven caffeoylquinic acids and their derivatives and five flavonoid glycosides. The production of caffeoylquinic acids and their derivatives was elevated in hairy roots grown in the light. Only light-grown hairy roots demonstrated the capability for the biosynthesis of such flavonoid glycosides as quercetagetin, quercetin, luteolin, and patuletin hexosides. Chlorogenic acid, 3,5-di-O-caffeoylquinic acid and a tentatively identified tricaffeoylquinic acid derivative were detected as the major compounds present in the transformed roots. PMID:25811023

Bioactivity of Ruta graveolens and Satureja montana Essential Oils on Solanum tuberosum Hairy Roots and Solanum tuberosum Hairy Roots with Meloidogyne chitwoodi Co-cultures.

PubMed

Faria, Jorge M S; Rodrigues, Ana M; Sena, Inês; Moiteiro, Cristina; Bennett, Richard N; Mota, Manuel; Figueiredo, A Cristina

2016-10-12

As a nematotoxics screening biotechnological system, Solanum tuberosum hairy roots (StHR) and S. tuberosum hairy roots with Meloidogyne chitwoodi co-cultures (StHR/CRKN) were evaluated, with and without the addition of the essential oils (EOs) of Satureja montana and Ruta graveolens. EOs nematotoxic and phytotoxic effects were followed weekly by evaluating nematode population density in the co-cultures as well as growth and volatile profiles of both in vitro cultures types. Growth, measured by the dissimilation method and by fresh and dry weight determination, was inhibited after EO addition. Nematode population increased in control cultures, while in EO-added cultures numbers were kept stable. In addition to each of the EOs main components, and in vitro cultures constitutive volatiles, new volatiles were detected by gas chromatography and gas chromatography coupled to mass spectrometry in both culture types. StHR with CRKN co-cultures showed to be suitable for preliminary assessment of nematotoxic EOs.

Curcumin: a promising agent targeting cancer stem cells.

PubMed

Zang, Shufei; Liu, Tao; Shi, Junping; Qiao, Liang

2014-01-01

Cancer stem cells are a subset of cells that are responsible for cancer initiation and relapse. They are generally resistant to the current anticancer agents. Successful anticancer therapy must consist of approaches that can target not only the differentiated cancer cells, but also cancer stem cells. Emerging evidence suggested that the dietary agent curcumin exerted its anti-cancer activities via targeting cancer stem cells of various origins such as those of colorectal cancer, pancreatic cancer, breast cancer, brain cancer, and head and neck cancer. In order to enhance the therapeutic potential of curcumin, this agent has been modified or used in combination with other agents in the experimental therapy for many cancers. In this mini-review, we discussed the effect of curcumin and its derivatives in eliminating cancer stem cells and the possible underlying mechanisms.

Production of oleanolic acid glycosides by hairy root established cultures of Calendula officinalis L.

PubMed

Długosz, Marek; Wiktorowska, Ewa; Wiśniewska, Anita; Pączkowski, Cezary

2013-01-01

In order to initiate hairy root culture initiation cotyledons and hypocotyls of Calendula officinalis L. were infected with Agrobacterium rhizogenes strain ATCC 15834 or the same strain containing pCAMBIA 1381Z vector with β-glucuronidase reporter gene under control of promoter of NIK (Nematode Induced Kinase) gene. The efficiency of induction of hairy roots reached 33.8% for cotyledons and 66.6% for hypocotyls together for both transformation experiments. Finally, eight control and nine modified lines were established as a long-term culture. The hairy root cultures showed the ability to synthesize oleanolic acid mainly (97%) as glycosides; control lines contained it at the average 8.42 mg · g(-1) dry weight in tissue and 0.23 mg · dm(-3) in medium; modified lines: 4.59 mg · g(-1) for the tissue, and 0.48 mg · dm(-3) for the medium. Additionally lines showed high positive correlation between dry/fresh weight and oleanolic acid concentration in tissue. Using the Killiani mixture in acidic hydrolysis of oleanolic acid glycosides released free aglycones that were partially acetylated in such conditions.

Enhancement of Chlorogenic Acid Production in Hairy Roots of Platycodon grandiflorum by Over-Expression of An Arabidopsis thaliana Transcription Factor AtPAP1

PubMed Central

Tuan, Pham Anh; Kwon, Do Yeon; Lee, Sanghyun; Arasu, Mariadhas Valan; Al-Dhabi, Naif Abdullah; Park, Nam Il; Park, Sang Un

2014-01-01

To improve the production of chlorogenic acid (CGA) in hairy roots of Platycodon grandiflorum, we induced over-expression of Arabidopsis thaliana transcription factor production of anthocyanin pigment (AtPAP1) using an Agrobacterium rhizogenes-mediated transformation system. Twelve hairy root lines showing over-expression of AtPAP1 were generated. In order to investigate the regulation of AtPAP1 on the activities of CGA biosynthetic genes, the expression levels of seven P. grandiflorum CGA biosynthetic genes were analyzed in the hairy root line that had the greatest accumulation of AtPAP1 transcript, OxPAP1-1. The introduction of AtPAP1 increased the mRNA levels of all examined CGA biosynthetic genes and resulted in a 900% up-regulation of CGA accumulation in OxPAP1-1 hairy roots relative to controls. This suggests that P. grandiflorum hairy roots that over-express the AtPAP1 gene are a potential alternative source of roots for the production of CGA. PMID:25153629

Noncoherent-intense-pulsed light for the treatment of relapsing hairy intradermal melanocytic nevus after shave excision.

PubMed

Moreno-Arias, G A; Ferrando, J

2001-01-01

Few reports about melanocytic lesions treatment by means of noncoherent-intense-pulsed light (NCIPL) have been published. Here we evaluate the clinical results of a relapsing hairy intradermal melanocytic nevus treated with a noncoherent-intense-pulsed light source. A facial repigmented hairy intradermal melanocytic nevus that relapsed after shave excision, received four treatment sessions of a noncoherent-intense-pulsed light source (EpiLight, ESC Medical Systems Ltd, Israel) with the following parameters: 755 nm, a fluence energy of 40-42.5 J/cm(2), triple mode, a pulse width of 3.8 ms, and a delay of 20 ms, at 4-week intervals. Complete pigment clearance and hair removal was obtained. We have neither observed repigmentation nor hair regrowth after a 6 month-follow-up. No side effects were documented. Noncoherent-intense-pulse light is an effective treatment for hairy-pigmented melanocytic nevus. Copyright 2001 Wiley-Liss, Inc.

Effect of harvest timing and leaf hairiness on fiber quality

USDA-ARS?s Scientific Manuscript database

Recent concerns over leaf grades have generated questions of how both time of day cotton is harvested, as well as leaf hairiness levels of certain varieties, influence fiber quality. To address this, two smooth leaf varieties and two varieties with higher levels of leaf pubescence were harvested at...

On a Class of Hairy Square Barriers and Gamow Vectors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fernandez-Garcia, N.

The second order Darboux-Gamow transformation is applied to deform square one dimensional barriers in non-relativistic quantum mechanics. The initial and the new 'hairy' potentials have the same transmission probabilities (for the appropriate parameters). In general, new Gamow vectors are constructed as Darboux deformations of the initial ones.

Lytic agents, cell permeability, and monolayer penetrability.

PubMed

Salton, M R

1968-07-01

Cell lysis induced by lytic agents is the terminal phase of a series of events leading to membrane disorganization and breadkdown with the release of cellular macromolecules. Permeability changes following exposure to lytic systems may range from selective effects on ion fluxes to gross membrane damage and cell leakage. Lysis can be conceived as an interfacial phenomenon, and the action of surface-active agents on erythrocytes has provided a model in which to investigate relationships between hemolysis and chemical structure, ionic charge, surface tension lowering, and ability to penetrate monolayers of membrane lipid components. Evidence suggests that lysis follows the attainment of surface pressures exceeding a "critical collapse" level and could involve membrane cholesterol or phospholipid. Similarities of chemical composition of membranes from various cell types could account for lytic responses observed on interaction with surface-active agents. Cell membranes usually contain about 20-30 % lipid and 50-75 % protein. One or two major phospholipids are present in all cell membranes, but sterols are not detectable in bacterial membranes other than those of the Mycoplasma group. The rigid cell wall in bacteria has an important bearing on their response to treatment with lytic agents. Removal of the wall renders the protoplast membrane sensitive to rapid lysis with surfactants. Isolated membranes of erythrocytes and bacteria are rapidly dissociated by surface-active agents. Products of dissociation of bacterial membranes have uniform behavior in the ultracentrifuge (sedimentation coefficients 2-3S). Dissociation of membrane proteins from lipids and the isolation and characterization of these proteins will provide a basis for investigating the specificity of interaction of lytic agents with biomembranes.

Droplet evaporation and spread on waxy and hairy leaves associated with type and concentration of adjuvants.

PubMed

Xu, Linyun; Zhu, Heping; Ozkan, H Erdal; Bagley, William E; Krause, Charles R

2011-07-01

Adjuvants can improve pesticide application efficiency and effectiveness. However, quantifications of the adjuvant-amended pesticide droplet actions on foliage, which could affect application efficiencies, are largely unknown. Droplet evaporation rates and spread on waxy or hairy leaves varied greatly with the adjuvant types tested. On waxy leaves, the wetted areas of droplets containing crop oil concentrate (COC) were significantly smaller than those containing modified seed oil (MSO), non-ionic surfactant (NIS) or oil surfactant blend (OSB), whereas the evaporation rates of COC-amended droplets were significantly higher. On hairy leaves, COC-amended droplets remained on top of the hairs without wetting the epidermis. When the relative concentration was 1.50, the wetted area of droplets with NIS was 9.2 times lower than that with MSO and 6.1 times lower than that with OSB. The wetted area increased as the adjuvant concentration increased. MSO- or OSB-amended droplets spread extensively on the hairy leaf surface until they were completely dried. These results demonstrated that the proper concentration of MSO, NIS or OSB in spray mixtures improved the homogeneity of spray coverage on both waxy and hairy leaf surfaces and could reduce pesticide use. This article is a US Government work and is in the public domain in the USA. Published 2011 by John Wiley & Sons, Ltd.

Effect of Different Agrobacterium rhizogenes Strains on Hairy Root Induction and Phenylpropanoid Biosynthesis in Tartary Buckwheat (Fagopyrum tataricum Gaertn)

PubMed Central

Thwe, Aye; Valan Arasu, Mariadhas; Li, Xiaohua; Park, Chang Ha; Kim, Sun Ju; Al-Dhabi, Naif Abdullah; Park, Sang Un

2016-01-01

The development of an efficient protocol for successful hairy root induction by Agrobacterium rhizogenes is the key step toward an in vitro culturing method for the mass production of secondary metabolites. The selection of an effective Agrobacterium strain for the production of hairy roots is highly plant species dependent and must be determined empirically. Therefore, our goal was to investigate the transformation efficiency of different A. rhizogenes strains for the induction of transgenic hairy roots in Fagopyrum tataricum ‘Hokkai T10’ cultivar; to determine the expression levels of the polypropanoid biosynthetic pathway genes, such as ftpAL, FtC4H, Ft4CL, FrCHS, FrCH1, FrF3H, FtFLS1, FtFLS2, FtF3, H1, FtF3′H2, FtANS, and FtDFR; and to quantify the in vitro synthesis of phenolic compounds and anthocyanins. Among different strains, R1000 was the most promising candidate for hairy root stimulation because it induced the highest growth rate, root number, root length, transformation efficiency, and total anthocyanin and rutin content. The R1000, 15834, and A4 strains provided higher transcript levels for most metabolic pathway genes for the synthesis of rutin (22.31, 15.48, and 13.04 μg/mg DW, respectively), cyanidin 3-O-glucoside (800, 750, and 650 μg/g DW, respectively), and cyanidin 3-O-rutinoside (2410, 1530, and 1170 μg/g DW, respectively). A suitable A. rhizogenes strain could play a vital role in the fast growth of the bulk amount of hairy roots and secondary metabolites. Overall, R1000 was the most promising strain for hairy root induction in buckwheat. PMID:27014239

Hairy vetch seedbank persistence and implications for cover crop management

USDA-ARS?s Scientific Manuscript database

Hairy vetch (Vicia villosa Roth) is a fast growing, winter hardy annual legume that can produce shoot biomass levels upwards of 6500 kg ha-1. This cover crop is well suited for summer annual grain rotations, as it fixes considerable amounts of nitrogen, reduces erosion through rapid ground cover, an...

Scalar solitons and the microscopic entropy of hairy black holes in three dimensions

NASA Astrophysics Data System (ADS)

Correa, Francisco; Martínez, Cristián; Troncoso, Ricardo

2011-01-01

General Relativity coupled to a self-interacting scalar field in three dimensions is shown to admit exact analytic soliton solutions, such that the metric and the scalar field are regular everywhere. Since the scalar field acquires slow fall-off at infinity, the soliton describes an asymptotically AdS spacetime in a relaxed sense as compared with the one of Brown and Henneaux. Nevertheless, the asymptotic symmetry group remains to be the conformal group, and the algebra of the canonical generators possesses the standard central extension. For this class of asymptotic behavior, the theory also admits hairy black holes which raises some puzzles concerning a holographic derivation of their entropy à la Strominger. Since the soliton is devoid of integration constants, it has a fixed (negative) mass, and it can be naturally regarded as the ground state of the "hairy sector", for which the scalar field is switched on. This assumption allows to exactly reproduce the semiclassical hairy black hole entropy from the asymptotic growth of the number of states by means of Cardy formula. Particularly useful is expressing the asymptotic growth of the number of states only in terms of the spectrum of the Virasoro operators without making any explicit reference to the central charges.

Linking a Germplasm Collection of the Cover Crop Hairy Vetch (Vicia villosa Roth) to Traits Related to Improved Nitrogen Fixation

USDA-ARS?s Scientific Manuscript database

Hairy vetch is used as a leguminous cover crop throughout the United States providing important ecosystem services in agro-ecosystems (Abdul-Baki et al., 2002; Mohler and Teasdale, 1993; Puget and Drinkwater, 2001; Seo et al., 2006; Stute and Posner, 1995). Many traits found in hairy vetch have pro...

Thermoreceptive innervation of human glabrous and hairy skin: a contact heat evoked potential analysis.

PubMed

Granovsky, Yelena; Matre, Dagfinn; Sokolik, Alexander; Lorenz, Jürgen; Casey, Kenneth L

2005-06-01

The human palm has a lower heat detection threshold and a higher heat pain threshold than hairy skin. Neurophysiological studies of monkeys suggest that glabrous skin has fewer low threshold heat nociceptors (AMH type 2) than hairy skin. Accordingly, we used a temperature-controlled contact heat evoked potential (CHEP) stimulator to excite selectively heat receptors with C fibers or Adelta-innervated AMH type 2 receptors in humans. On the dorsal hand, 51 degrees C stimulation produced painful pinprick sensations and 41 degrees C stimuli evoked warmth. On the glabrous thenar, 41 degrees C stimulation produced mild warmth and 51 degrees C evoked strong but painless heat sensations. We used CHEP responses to estimate the conduction velocities (CV) of peripheral fibers mediating these sensations. On hairy skin, 41 degrees C stimuli evoked an ultra-late potential (mean, SD; N wave latency: 455 (118) ms) mediated by C fibers (CV by regression analysis: 1.28 m/s, N=15) whereas 51 degrees C stimuli evoked a late potential (N latency: 267 (33) ms) mediated by Adelta afferents (CV by within-subject analysis: 12.9 m/s, N=6). In contrast, thenar responses to 41 and 51 degrees C were mediated by C fibers (average N wave latencies 485 (100) and 433 (73) ms, respectively; CVs 0.95-1.35 m/s by regression analysis, N=15; average CV=1.7 (0.41) m/s calculated from distal glabrous and proximal hairy skin stimulation, N=6). The exploratory range of the human and monkey palm is enhanced by the abundance of low threshold, C-innervated heat receptors and the paucity of low threshold AMH type 2 heat nociceptors.

Superradiant instability of near extremal and extremal four-dimensional charged hairy black holes in anti-de Sitter spacetime

NASA Astrophysics Data System (ADS)

González, P. A.; Papantonopoulos, Eleftherios; Saavedra, Joel; Vásquez, Yerko

2017-03-01

We study the instability of near extremal and extremal four-dimensional anti-de Sitter charged hairy black holes to radial neutral massive and charged massless scalar field perturbations. We solve the scalar field equation by using the improved asymptotic iteration method and the time domain analysis, and we find the quasinormal frequencies. For the charged scalar perturbations, we find the superradiance condition by computing the reflection coefficient in the low-frequency limit, and we show that in the superradiance regime, which depends on the scalar hair charge, all modes of radial charged massless perturbations are unstable, indicating that the charged hairy black hole is superradiantly unstable. On the other hand, calculating the quasinormal frequencies of radial neutral scalar perturbations in this background, we find stability of the charged hairy black hole.

Modelling of robotic work cells using agent based-approach

NASA Astrophysics Data System (ADS)

Sękala, A.; Banaś, W.; Gwiazda, A.; Monica, Z.; Kost, G.; Hryniewicz, P.

2016-08-01

In the case of modern manufacturing systems the requirements, both according the scope and according characteristics of technical procedures are dynamically changing. This results in production system organization inability to keep up with changes in a market demand. Accordingly, there is a need for new design methods, characterized, on the one hand with a high efficiency and on the other with the adequate level of the generated organizational solutions. One of the tools that could be used for this purpose is the concept of agent systems. These systems are the tools of artificial intelligence. They allow assigning to agents the proper domains of procedures and knowledge so that they represent in a self-organizing system of an agent environment, components of a real system. The agent-based system for modelling robotic work cell should be designed taking into consideration many limitations considered with the characteristic of this production unit. It is possible to distinguish some grouped of structural components that constitute such a system. This confirms the structural complexity of a work cell as a specific production system. So it is necessary to develop agents depicting various aspects of the work cell structure. The main groups of agents that are used to model a robotic work cell should at least include next pattern representatives: machine tool agents, auxiliary equipment agents, robots agents, transport equipment agents, organizational agents as well as data and knowledge bases agents. In this way it is possible to create the holarchy of the agent-based system.

Production of ajmalicine and ajmaline in hairy root cultures of Rauvolfia micrantha Hook f., a rare and endemic medicinal plant.

PubMed

Sudha, C G; Obul Reddy, B; Ravishankar, G A; Seeni, S

2003-04-01

Hairy roots of Rauvolfia micrantha were induced from hypocotyl explants of 2-3 weeks old aseptic seedlings using Agrobacterium rhizogenes ATCC 15834. Hairy roots grown in half-strength Murashige & Skoog (MS) medium with 0.2 mg indole 3-butyric acid l-1 and 0.1 mg alpha-naphthaleneacetic acid l-1 produced more ajmaline (0.01 mg g-1 dry wt) and ajmalicine (0.006 mg g-1 dry wt) than roots grown in auxin-free medium. Ajmaline (0.003 mg g-1 dry wt) and ajmalicine (0.0007 mg g-1 dry wt) were also produced in normal root cultures. This is the first report of production of ajmaline and ajmalicine in hairy root cultures of Rauvolfia micrantha.

The role of lin-22, a hairy/enhancer of split homolog, in patterning the peripheral nervous system of C. elegans.

PubMed

Wrischnik, L A; Kenyon, C J

1997-08-01

In C. elegans, six lateral epidermal stem cells, the seam cells V1-V6, are located in a row along the anterior-posterior (A/P) body axis. Anterior seam cells (V1-V4) undergo a fairly simple sequence of stem cell divisions and generate only epidermal cells. Posterior seam cells (V5 and V6) undergo a more complicated sequence of cell divisions that include additional rounds of stem cell proliferation and the production of neural as well as epidermal cells. In the wild type, activity of the gene lin-22 allows V1-V4 to generate their normal epidermal lineages rather than V5-like lineages. lin-22 activity is also required to prevent additional neurons from being produced by one branch of the V5 lineage. We find that the lin-22 gene exhibits homology to the Drosophila gene hairy, and that lin-22 activity represses neural development within the V5 lineage by blocking expression of the posterior-specific Hox gene mab-5 in specific cells. In addition, in order to prevent anterior V cells from generating V5-like lineages, wild-type lin-22 gene activity must inhibit (directly or indirectly) at least five downstream regulatory gene activities. In anterior body regions, lin-22(+) inhibits expression of the Hox gene mab-5. It also inhibits the activity of the achaete-scute homolog lin-32 and an unidentified gene that we postulate regulates stem cell division. Each of these three genes is required for the expression of a different piece of the ectopic V5-like lineages generated in lin-22 mutants. In addition, lin-22 activity prevents two other Hox genes, lin-39 and egl-5, from acquiring new activities within their normal domains of function along the A/P body axis. Some, but not all, of the patterning activities of lin-22 in C. elegans resemble those of hairy in Drosophila.

Expression of important pathway genes involved in withanolides biosynthesis in hairy root culture of Withania somnifera upon treatment with Gracilaria edulis and Sargassum wightii.

PubMed

Sivanandhan, Ganeshan; Arunachalam, Chinnathambi; Selvaraj, Natesan; Sulaiman, Ali Alharbi; Lim, Yong Pyo; Ganapathi, Andy

2015-06-01

The investigation of seaweeds, Gracilaria edulis and Sargassum wightii extracts was carried out for the estimation of growth characteristics and major withanolides production in hairy root culture of Withania somnifera. The extract of G. edulis (50%) in MS liquid basal medium enabled maximum production of dry biomass (5.46 g DW) and withanolides contents (withanolide A 5.23 mg/g DW; withaferin A 2.24 mg/g DW and withanone 4.83 mg/g DW) in hairy roots after 40 days of culture with 48 h contact time. The obtained withanolides contents were significantly higher (2.32-fold-2.66-fold) in hairy root culture when compared to the control. RT PCR analysis of important pathway genes such as SE, SS, HMGR and FPPS exhibited substantial higher expression upon the seaweed extracts treatment in hairy root culture. This experiment would paw a platform for withanolides production in hairy root culture with the influence of sea weed extracts for pharmaceutical companies in the future. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

A hairy-leaf gene, BLANKET LEAF, of wild Oryza nivara increases photosynthetic water use efficiency in rice.

PubMed

Hamaoka, Norimitsu; Yasui, Hideshi; Yamagata, Yoshiyuki; Inoue, Yoko; Furuya, Naruto; Araki, Takuya; Ueno, Osamu; Yoshimura, Atsushi

2017-12-01

High water use efficiency is essential to water-saving cropping. Morphological traits that affect photosynthetic water use efficiency are not well known. We examined whether leaf hairiness improves photosynthetic water use efficiency in rice. A chromosome segment introgression line (IL-hairy) of wild Oryza nivara (Acc. IRGC105715) with the genetic background of Oryza sativa cultivar 'IR24' had high leaf pubescence (hair). The leaf hairs developed along small vascular bundles. Linkage analysis in BC 5 F 2 and F 3 populations showed that the trait was governed by a single gene, designated BLANKET LEAF (BKL), on chromosome 6. IL-hairy plants had a warmer leaf surface in sunlight, probably due to increased boundary layer resistance. They had a lower transpiration rate under moderate and high light intensities, resulting in higher photosynthetic water use efficiency. Introgression of BKL on chromosome 6 from O. nivara improved photosynthetic water use efficiency in the genetic background of IR24.

Dynamical formation of a hairy black hole in a cavity from the decay of unstable solitons

NASA Astrophysics Data System (ADS)

Sanchis-Gual, Nicolas; Degollado, Juan Carlos; Font, José A.; Herdeiro, Carlos; Radu, Eugen

2017-08-01

Recent numerical relativity simulations within the Einstein-Maxwell-(charged-)Klein-Gordon (EMcKG) system have shown that the non-linear evolution of a superradiantly unstable Reissner-Nordström black hole (BH) enclosed in a cavity, leads to the formation of a BH with scalar hair. Perturbative evidence for the stability of such hairy BHs has been independently established, confirming they are the true endpoints of superradiant instability. The same EMcKG system admits also charged scalar soliton-type solutions, which can be either stable or unstable. Using numerical relativity techniques, we provide evidence that the time evolution of some of these unstable solitons leads, again, to the formation of a hairy BH. In some other cases, unstable solitons evolve into a (bald) Reissner-Nordström BH. These results establish that the system admits two distinct channels to form hairy BHs at the threshold of superradiance: growing hair from an unstable (bald) BH, or growing a horizon from an unstable (horizonless) soliton. Some parallelism with the case of asymptotically flat boson stars and Kerr BHs with scalar hair is drawn.

Equine Airway Mast Cells are Sensitive to Cell Death Induced by Lysosomotropic Agents.

PubMed

Wernersson, S; Riihimäki, M; Pejler, G; Waern, I

2017-01-01

Mast cells are known for their detrimental effects in various inflammatory conditions. Regimens that induce selective mast cell apoptosis may therefore be of therapeutic significance. Earlier studies have demonstrated that murine- and human-cultured mast cells are highly sensitive to apoptosis induced by the lysosomotropic agent LeuLeuOMe (LLME). However, the efficacy of lysosomotropic agents for inducing apoptosis of in vivo-derived airway mast cells and the impact on mast cells in other species have not been assessed. Here we addressed whether lysosomotropic agents can induce cell death of equine in vivo-derived mast cells. Bronchoalveolar lavage (BAL) fluids from horses were incubated with LLME at 15-100 μm for up to 48 h. The overall cell viability was unaffected by 15 μm LLME up to 48 h, whereas a relatively modest drop in total cell counts (~30%) was seen at the highest LLME dose used. In contrast to the relatively low effect on total cell counts, LLME efficiently and dose dependently reduced the number of mast cells in BAL fluids, with an almost complete depletion (96%) of mast cells after 24 h of incubation with 100 μm LLME. A significant but less dramatic reduction (up to ~45%) of lymphocytes was also seen, whereas macrophages and neutrophils were essentially resistant. The appearance of apoptotic bodies suggested a mechanism involving apoptosis rather than necrosis. These findings suggest that equine airway mast cells are highly sensitive to lysosomotropic agents. Possibly, lysosomotropic agents could be of therapeutic value to treat disorders involving harmful accumulation of mast cells in the airways. © 2016 The Foundation for the Scandinavian Journal of Immunology.

Soft hairy warped black hole entropy

NASA Astrophysics Data System (ADS)

Grumiller, Daniel; Hacker, Philip; Merbis, Wout

2018-02-01

We reconsider warped black hole solutions in topologically massive gravity and find novel boundary conditions that allow for soft hairy excitations on the horizon. To compute the associated symmetry algebra we develop a general framework to compute asymptotic symmetries in any Chern-Simons-like theory of gravity. We use this to show that the near horizon symmetry algebra consists of two u (1) current algebras and recover the surprisingly simple entropy formula S = 2 π( J 0 + + J 0 - ), where J 0 ± are zero mode charges of the current algebras. This provides the first example of a locally non-maximally symmetric configuration exhibiting this entropy law and thus non-trivial evidence for its universality.

Biodegradation of γ-hexachlorocyclohexane by transgenic hairy root cultures of Cucurbita moschata that accumulate recombinant bacterial LinA.

PubMed

Nanasato, Yoshihiko; Namiki, Sayuri; Oshima, Masao; Moriuchi, Ryota; Konagaya, Ken-Ichi; Seike, Nobuyasu; Otani, Takashi; Nagata, Yuji; Tsuda, Masataka; Tabei, Yutaka

2016-09-01

γ-HCH was successfully degraded using LinA-expressed transgenic hairy root cultures of Cucurbita moschata . Fusing an endoplasmic reticulum-targeting signal peptide to LinA was essential for stable accumulation in the hairy roots. The pesticide γ-hexachlorocyclohexane (γ-HCH) is a persistent organic pollutant (POP) that raises public health and environmental pollution concerns worldwide. Although several isolates of γ-HCH-degrading bacteria are available, inoculating them directly into γ-HCH-contaminated soil is ineffective because of the bacterial survival rate. Cucurbita species incorporate significant amounts of POPs from soils compared with other plant species. Here, we describe a novel bioremediation strategy that combines the bacterial degradation of γ-HCH and the efficient uptake of γ-HCH by Cucurbita species. We produced transgenic hairy root cultures of Cucurbita moschata that expressed recombinant bacterial linA, isolated from the bacterium Sphingobium japonicum UT26. The LinA protein was accumulated stably in the hairy root cultures by fusing an endoplasmic reticulum (ER)-targeting signal peptide to LinA. Then, we demonstrated that the cultures degraded more than 90 % of γ-HCH (1 ppm) overnight and produced the γ-HCH metabolite 1,2,4-trichlorobenzene, indicating that LinA degraded γ-HCH. These results indicate that the gene linA has high potential for phytoremediation of environmental γ-HCH.

Influence of Seeding Ratio, Planting Date, and Termination Date on Rye-Hairy Vetch Cover Crop Mixture Performance under Organic Management

PubMed Central

Lawson, Andrew; Cogger, Craig; Bary, Andy; Fortuna, Ann-Marie

2015-01-01

Cover crop benefits include nitrogen accumulation and retention, weed suppression, organic matter maintenance, and reduced erosion. Organic farmers need region-specific information on winter cover crop performance to effectively integrate cover crops into their crop rotations. Our research objective was to compare cover crop seeding mixtures, planting dates, and termination dates on performance of rye (Secale cereale L.) and hairy vetch (Vicia villosa Roth) monocultures and mixtures in the maritime Pacific Northwest USA. The study included four seed mixtures (100% hairy vetch, 25% rye-75% hairy vetch, 50% rye-50% hairy vetch, and 100% rye by seed weight), two planting dates, and two termination dates, using a split-split plot design with four replications over six years. Measurements included winter ground cover; stand composition; cover crop biomass, N concentration, and N uptake; and June soil NO3 --N. Rye planted in mid-September and terminated in late April averaged 5.1 Mg ha-1 biomass, whereas mixtures averaged 4.1 Mg ha-1 and hairy vetch 2.3 Mg ha-1. Delaying planting by 2.5 weeks reduced average winter ground cover by 65%, biomass by 50%, and cover crop N accumulation by 40%. Similar reductions in biomass and N accumulation occurred for late March termination, compared with late April termination. Mixtures had less annual biomass variability than rye. Mixtures accumulated 103 kg ha-1 N and had mean C:N ratio <17:1 when planted in mid-September and terminated in late April. June soil NO3 --N (0 to 30 cm depth) averaged 62 kg ha-1 for rye, 97 kg ha-1 for the mixtures, and 119 kg ha-1 for hairy vetch. Weeds comprised less of the mixtures biomass (20% weeds by weight at termination) compared with the monocultures (29%). Cover crop mixtures provided a balance between biomass accumulation and N concentration, more consistent biomass over the six-year study, and were more effective at reducing winter weeds compared with monocultures. PMID:26080008

Influence of Seeding Ratio, Planting Date, and Termination Date on Rye-Hairy Vetch Cover Crop Mixture Performance under Organic Management.

PubMed

Lawson, Andrew; Cogger, Craig; Bary, Andy; Fortuna, Ann-Marie

2015-01-01

Cover crop benefits include nitrogen accumulation and retention, weed suppression, organic matter maintenance, and reduced erosion. Organic farmers need region-specific information on winter cover crop performance to effectively integrate cover crops into their crop rotations. Our research objective was to compare cover crop seeding mixtures, planting dates, and termination dates on performance of rye (Secale cereale L.) and hairy vetch (Vicia villosa Roth) monocultures and mixtures in the maritime Pacific Northwest USA. The study included four seed mixtures (100% hairy vetch, 25% rye-75% hairy vetch, 50% rye-50% hairy vetch, and 100% rye by seed weight), two planting dates, and two termination dates, using a split-split plot design with four replications over six years. Measurements included winter ground cover; stand composition; cover crop biomass, N concentration, and N uptake; and June soil NO3(-)-N. Rye planted in mid-September and terminated in late April averaged 5.1 Mg ha(-1) biomass, whereas mixtures averaged 4.1 Mg ha(-1) and hairy vetch 2.3 Mg ha(-1). Delaying planting by 2.5 weeks reduced average winter ground cover by 65%, biomass by 50%, and cover crop N accumulation by 40%. Similar reductions in biomass and N accumulation occurred for late March termination, compared with late April termination. Mixtures had less annual biomass variability than rye. Mixtures accumulated 103 kg ha(-1) N and had mean C:N ratio <17:1 when planted in mid-September and terminated in late April. June soil NO3(-)-N (0 to 30 cm depth) averaged 62 kg ha(-1) for rye, 97 kg ha(-1) for the mixtures, and 119 kg ha(-1) for hairy vetch. Weeds comprised less of the mixtures biomass (20% weeds by weight at termination) compared with the monocultures (29%). Cover crop mixtures provided a balance between biomass accumulation and N concentration, more consistent biomass over the six-year study, and were more effective at reducing winter weeds compared with monocultures.

Spatial temperature distribution in human hairy and glabrous skin after infrared CO2 laser radiation

PubMed Central

2010-01-01

Background CO2 lasers have been used for several decades as an experimental non-touching pain stimulator. The laser energy is absorbed by the water content in the most superficial layers of the skin. The deeper located nociceptors are activated by passive conduction of heat from superficial to deeper skin layers. Methods In the current study, a 2D axial finite element model was developed and validated to describe the spatial temperature distribution in the skin after infrared CO2 laser stimulation. The geometry of the model was based on high resolution ultrasound scans. The simulations were compared to the subjective pain intensity ratings from 16 subjects and to the surface skin temperature distributions measured by an infrared camera. Results The stimulations were sensed significantly slower and less intense in glabrous skin than they were in hairy skin (MANOVA, p < 0.001). The model simulations of superficial temperature correlated with the measured skin surface temperature (r > 0.90, p < 0.001). Of the 16 subjects tested; eight subjects reported pricking pain in the hairy skin following a stimulus of 0.6 J/cm2 (5 W, 0.12 s, d1/e2 = 11.4 mm) only two reported pain to glabrous skin stimulation using the same stimulus intensity. The temperature at the epidermal-dermal junction (depth 50 μm in hairy and depth 133 μm in glabrous skin) was estimated to 46°C for hairy skin stimulation and 39°C for glabrous skin stimulation. Conclusions As compared to previous one dimensional heat distribution models, the current two dimensional model provides new possibilities for detailed studies regarding CO2 laser stimulation intensity, temperature levels and nociceptor activation. PMID:21059226

Nootropic agents stimulate neurogenesis. Brain Cells, Inc.: WO2007104035.

PubMed

Taupin, Philippe

2009-05-01

The application is in the field of adult neurogenesis, neural stem cells and cellular therapy. It aims to characterize the activity of nootropic agents on adult neurogenesis in vitro. Nootropic agents are substances improving cognitive and mental abilities. AMPA (alpha-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate) and nootropic agents were assessed for the potential to differentiate human neural progenitor and stem cells into neuronal cells in vitro. They were also tested for their behavioural activity on the novel object recognition task. AMPA, piracetam, FK-960 and SGS-111 induce and stimulate neuronal differentiation of human-derived neural progenitor and stem cells. SGS-111 increases the number of visits to the novel object. The neurogenic activity of piracetam and SGS-111 is mediated through AMPA receptor. The neurogenic activity of SGS-111 may contribute and play a role in its nootropic activity. These results suggest that nootropic agents may elicit some of their effects through their neurogenic activity. The application claims the use of nootropic agents for their neurogenic activity and for the treatment of neurological diseases, disorders and injuries, by stimulating or increasing the generation of neuronal cells in the adult brain.

Enhanced production of azadirachtin by hairy root cultures of Azadirachta indica A. Juss by elicitation and media optimization.

PubMed

Satdive, Ramesh K; Fulzele, Devanand P; Eapen, Susan

2007-02-01

Azadirachtin is one of the most potent biopesticides so far developed from a plant sources. Influence of different culture media and elicitation on growth and production of azadirachtin by hairy root cultures of Azadirachta indica was studied. Out of the three media tested, namely Ohyama and Nitsch, Gamborg's and Murashige and Skoog's basal media, hairy roots cultured on Ohyama and Nitsch's basal medium produced maximum yield of azadirachtin (0.0166% dry weight, DW). Addition of biotic elicitor enhanced the production of azadirachtin by approximately 5-fold (0.074% DW), while signal compounds such as jasmonic acid and salicylic acid showed a approximately 6 (0.095% DW) and approximately 9-fold (0.14% DW) enhancement, respectively, in the production of azadirachtin as compared to control cultures on Ohyama and Nitsch medium. Extracts from hairy roots were found to be superior to those from the leaves for antifeedant activity against the larvae of Spodoptera litura.

Benzamide Derivatives as Protective Agents against the Action of Xenotoxic Agents on Human Cells.

DTIC Science & Technology

1984-05-31

4D-AlI45 396 BENZAMIDE DERIVATIVES AS PROTECTIVE AGENTS AGAINST THE I/i ACTION OF XENOTOXI..(U) OHIO STATE UNIV RESEARCH I FOUNDATION COLUMBUS G E...AS PROTECTIVE AGENTS AGAINST THE ACTION OF XENOTOXIC AGENTS ON HUMAN CELLS CD George E. Milo * Department of Physiological Chemistry and...TITLE (and Subtitle) S. .YPE OF REPORT & PERIOD COVERED Benzamide Derivatives as Protective Agents Annual Scientific Report 5 Against the Action of

Genetic elicitation by inducible expression of β-cryptogein stimulates secretion of phenolics from Coleus blumei hairy roots.

PubMed

Vuković, Rosemary; Bauer, Nataša; Curković-Perica, Mirna

2013-02-01

The accumulation of phenolic compounds in plants is often part of the defense response against stress and pathogen attack, which can be triggered and activated by elicitors. Oomycetal proteinaceous elicitor, β-cryptogein, induces hypersensitive response and systemic acquired resistance against some pathogens. In order to test the effect of endogenously synthesized cryptogein protein on phenolic compounds accumulation in tissue, and secretion into the culture medium, Coleus blumei hairy roots were generated. Agrobacterium rhizogenes was employed to insert synthetic crypt gene, encoding β-cryptogein, under the control of alcohol-inducible promoter. The expression of β-cryptogein, in C. blumei hairy roots, was controlled by application of 1% and 2% ethanol, during 21 days induction period. Ethanol-induced expression of β-cryptogein caused significant decrease of soluble phenolics and rosmarinic acid (RA) in hairy root lines and increase of phenolics, RA and caffeic acid in culture medium. These data suggest that β-cryptogein might be a potential regulatory factor for phenolics secretion from the roots. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Factors influencing plant regeneration from seedling explants of Hairy nightshade (Solanum sarrachoides)

USDA-ARS?s Scientific Manuscript database

A good model plant to investigate plant – pathogen interactions would be easy to grow, have a short life cycle, be a natural host of many pathogens, and be easy to manipulate genetically. Hairy nightshade (Solanum sarrachoides) is a ubiquitous, fast growing weed that produces copious amounts of see...

Iron Oxide as an MRI Contrast Agent for Cell Tracking

PubMed Central

Korchinski, Daniel J.; Taha, May; Yang, Runze; Nathoo, Nabeela; Dunn, Jeff F.

2015-01-01

Iron oxide contrast agents have been combined with magnetic resonance imaging for cell tracking. In this review, we discuss coating properties and provide an overview of ex vivo and in vivo labeling of different cell types, including stem cells, red blood cells, and monocytes/macrophages. Furthermore, we provide examples of applications of cell tracking with iron contrast agents in stroke, multiple sclerosis, cancer, arteriovenous malformations, and aortic and cerebral aneurysms. Attempts at quantifying iron oxide concentrations and other vascular properties are examined. We advise on designing studies using iron contrast agents including methods for validation. PMID:26483609

Sporamin-mediated resistance to beet cyst nematodes (Heterodera schachtii Schm.) is dependent on trypsin inhibitory activity in sugar beet (Beta vulgaris L.) hairy roots.

PubMed

Cai, Daguang; Thurau, Tim; Tian, Yanyan; Lange, Tina; Yeh, Kai-Wun; Jung, Christian

2003-04-01

Sporamin, a sweet potato tuberous storage protein, is a Kunitz-type trypsin inhibitor. Its capability of conferring insect-resistance on transgenic tobacco and cauliflower has been confirmed. To test its potential as an anti-feedant for the beet cyst nematode (Heterodera schachtii Schm.), the sporamin gene SpTI-1 was introduced into sugar beet (Beta vulgaris L.) by Agrobacterium rhizogenes-mediated transformation. Twelve different hairy root clones expressing sporamin were selected for studying nematode development. Of these, 8 hairy root clones were found to show significant efficiency in inhibiting the growth and development of the female nematodes whereas 4 root clones did not show any inhibitory effects even though the SpTI-1 gene was regularly expressed in all of the tested hairy roots as revealed by northern and western analyses. Inhibition of nematode development correlated with trypsin inhibitor activity but not with the amount of sporamin expressed in hairy roots. These data demonstrate that the trypsin inhibitor activity is the critical factor for inhibiting growth and development of cyst nematodes in sugar beet hairy roots expressing the sporamin gene. Hence, the sweet potato sporamin can be used as a new and effective anti-feedant for controlling cyst nematodes offering an alternative strategy for establishing nematode resistance in crops.

Carbohydrate and elicitor enhanced withanolide (withaferin A and withanolide A) accumulation in hairy root cultures of Withania somnifera (L.).

PubMed

Doma, Madhavi; Abhayankar, Gauri; Reddy, V D; Kavi Kishor, P B

2012-07-01

Leaves of Withania somnifera contained more withaferin A and withanolide A than roots indicating that these compounds mainly accumulate in leaves. With an increase in age of the plant, withaferin A was enhanced with a corresponding decrease in withanolide A. Hairy root cultures were induced from leaf explants using Agrobacterium rhizogenes and the transgenic nature of hairy roots was confirmed by partial isolation and sequencing of rolB gene, which could not be amplified in untransformed plant parts. In hairy roots, withaferin A accumulated at 2, 3 and 4% but not at 6% sucrose, the highest amount being 1733 microg/g dry weight at 4% level. High and equal amounts of withaferin A and withanolide A accumulated (890 and 886 microg/g dry tissue respectively) only at 3% sucrose. Increasing concentrations of glucose enhanced withaferin A and it peaked at 5% level (3866 microg/g dry tissue). This amount is 2842 and 34% higher compared to untransformed roots and leaves (collected from 210-day-old plants) respectively. Withanolide A was detected at 5% glucose but not at other concentrations. While chitosan and nitric oxide increased withaferin A, jasmonic acid decreased it. Acetyl salicylic acid stimulated accumulation of both withaferin A and withanolide A at higher concentrations. Triadimefon, a fungicide, enhanced withaferin A by 1626 and 3061% (not detected earlier) compared to hairy and intact roots respectively.

Bubble Jet agent release cartridge for chemical single cell stimulation.

PubMed

Wangler, N; Welsche, M; Blazek, M; Blessing, M; Vervliet-Scheebaum, M; Reski, R; Müller, C; Reinecke, H; Steigert, J; Roth, G; Zengerle, R; Paust, N

2013-02-01

We present a new method for the distinct specific chemical stimulation of single cells and small cell clusters within their natural environment. By single-drop release of chemical agents with droplets in size of typical cell diameters (d <30 μm) on-demand micro gradients can be generated for the specific manipulation of single cells. A single channel and a double channel agent release cartridge with integrated fluidic structures and integrated agent reservoirs are shown, tested, and compared in this publication. The single channel setup features a fluidic structure fabricated by anisotropic etching of silicon. To allow for simultaneous release of different agents even though maintaining the same device size, the second type comprises a double channel fluidic structure, fabricated by photolithographic patterning of TMMF. Dispensed droplet volumes are V = 15 pl and V = 10 pl for the silicon and the TMMF based setups, respectively. Utilizing the agent release cartridges, the application in biological assays was demonstrated by hormone-stimulated premature bud formation in Physcomitrella patens and the individual staining of one single L 929 cell within a confluent grown cell culture.

Expression of rabies glycoprotein and ricin toxin B chain (RGP-RTB) fusion protein in tomato hairy roots: a step towards oral vaccination for rabies.

PubMed

Singh, Ankit; Srivastava, Subhi; Chouksey, Ankita; Panwar, Bhupendra Singh; Verma, Praveen C; Roy, Sribash; Singh, Pradhyumna K; Saxena, Gauri; Tuli, Rakesh

2015-04-01

Transgenic hairy roots of Solanum lycopersicum were engineered to express a recombinant protein containing a fusion of rabies glycoprotein and ricin toxin B chain (rgp-rtxB) antigen under the control of constitutive CaMV35S promoter. Asialofetuin-mediated direct ELISA of transgenic hairy root extracts was performed using polyclonal anti-rabies antibodies (Ab1) and epitope-specific peptidal anti-RGP (Ab2) antibodies which confirmed the expression of functionally viable RGP-RTB fusion protein. Direct ELISA based on asialofetuin-binding activity was used to screen crude protein extracts from five transgenic hairy root lines. Expressions of RGP-RTB fusion protein in different tomato hairy root lines varied between 1.4 and 8 µg in per gram of tissue. Immunoblotting assay of RGP-RTB fusion protein from these lines showed a protein band on monomeric size of ~84 kDa after denaturation. Tomato hairy root line H03 showed highest level of RGP-RTB protein expression (1.14 %) and was used further in bench-top bioreactor for the optimization of scale-up process to produce large quantity of recombinant protein. Partially purified RGP-RTB fusion protein was able to induce the immune response in BALB/c mice after intra-mucosal immunization. In the present investigation, we have not only successfully scaled up the hairy root culture but also established the utility of this system to produce vaccine antigen which subsequently will reduce the total production cost for implementing rabies vaccination programs in developing nations. This study in a way aims to provide consolidated base for low-cost preparation of improved oral vaccine against rabies.

Thyroid Hormone‐Induced Activation of Notch Signaling is Required for Adult Intestinal Stem Cell Development During Xenopus Laevis Metamorphosis

PubMed Central

Fujimoto, Kenta; Kajita, Mitsuko; Fu, Liezhen; Shi, Yun‐Bo; Ishizuya‐Oka, Atsuko

2016-01-01

Abstract In Xenopus laevis intestine during metamorphosis, the larval epithelial cells are removed by apoptosis, and the adult epithelial stem (AE) cells appear concomitantly. They proliferate and differentiate to form the adult epithelium (Ep). Thyroid hormone (TH) is well established to trigger this remodeling by regulating the expression of various genes including Notch receptor. To study the role of Notch signaling, we have analyzed the expression of its components, including the ligands (DLL and Jag), receptor (Notch), and targets (Hairy), in the metamorphosing intestine by real‐time reverse transcription‐polymerase chain reaction and in situ hybridization or immunohistochemistry. We show that they are up‐regulated during both natural and TH‐induced metamorphosis in a tissue‐specific manner. Particularly, Hairy1 is specifically expressed in the AE cells. Moreover, up‐regulation of Hairy1 and Hairy2b by TH was prevented by treating tadpoles with a γ‐secretase inhibitor (GSI), which inhibits Notch signaling. More importantly, TH‐induced up‐regulation of LGR5, an adult intestinal stem cell marker, was suppressed by GSI treatment. Our results suggest that Notch signaling plays a role in stem cell development by regulating the expression of Hairy genes during intestinal remodeling. Furthermore, we show with organ culture experiments that prolonged exposure of tadpole intestine to TH plus GSI leads to hyperplasia of secretory cells and reduction of absorptive cells. Our findings here thus provide evidence for evolutionarily conserved role of Notch signaling in intestinal cell fate determination but more importantly reveal, for the first time, an important role of Notch pathway in the formation of adult intestinal stem cells during vertebrate development. Stem Cells 2017;35:1028–1039 PMID:27870267

Hairy Root Transformation Using Agrobacterium rhizogenes as a Tool for Exploring Cell Type-Specific Gene Expression and Function Using Tomato as a Model1[W][OPEN

PubMed Central

Ron, Mily; Kajala, Kaisa; Pauluzzi, Germain; Wang, Dongxue; Reynoso, Mauricio A.; Zumstein, Kristina; Garcha, Jasmine; Winte, Sonja; Masson, Helen; Inagaki, Soichi; Federici, Fernán; Sinha, Neelima; Deal, Roger B.; Bailey-Serres, Julia; Brady, Siobhan M.

2014-01-01

Agrobacterium rhizogenes (or Rhizobium rhizogenes) is able to transform plant genomes and induce the production of hairy roots. We describe the use of A. rhizogenes in tomato (Solanum spp.) to rapidly assess gene expression and function. Gene expression of reporters is indistinguishable in plants transformed by Agrobacterium tumefaciens as compared with A. rhizogenes. A root cell type- and tissue-specific promoter resource has been generated for domesticated and wild tomato (Solanum lycopersicum and Solanum pennellii, respectively) using these approaches. Imaging of tomato roots using A. rhizogenes coupled with laser scanning confocal microscopy is facilitated by the use of a membrane-tagged protein fused to a red fluorescent protein marker present in binary vectors. Tomato-optimized isolation of nuclei tagged in specific cell types and translating ribosome affinity purification binary vectors were generated and used to monitor associated messenger RNA abundance or chromatin modification. Finally, transcriptional reporters, translational reporters, and clustered regularly interspaced short palindromic repeats-associated nuclease9 genome editing demonstrate that SHORT-ROOT and SCARECROW gene function is conserved between Arabidopsis (Arabidopsis thaliana) and tomato. PMID:24868032

Enhanced daidzin production from jasmonic and acetyl salicylic acid elicited hairy root cultures of Psoralea corylifolia L. (Fabaceae).

PubMed

Zaheer, Mohd; Reddy, Vudem Dashavantha; Giri, Charu Chandra

2016-07-01

Daidzin (7-O-glucoside of daidzein) has several pharmacological benefits in herbal remedy, as antioxidant and shown antidipsotropic activity. Hairy root culture of Psoralea corylifolia L. was developed for biomass and enhanced daidzin production using signalling compounds such as jasmonic acid (JA) and acetyl salicylic acid (ASA). Best response of 2.8-fold daidzin (5.09% DW) with 1 μM JA treatment after second week and 7.3-fold (3.43% DW) with 10 μM JA elicitation after 10th week was obtained from hairy roots compared to untreated control. ASA at 10 μM promoted 1.7-fold increase in daidzin (1.49% DW) content after seventh week compared to control (0.83% DW). Addition of 25 μM ASA resulted in 1.44% DW daidzin (1.5-fold increase) with 0.91% DW in control after fifth week and 1.44% DW daidzin (2.3-fold increase) after eighth week when compared to untreated control (0.62% DW). Reduced biomass with increased daidzin content was facilitated by elicited hairy root cultures.

Striga parasitizes transgenic hairy roots of Zea mays and provides a tool for studying plant-plant interactions

PubMed Central

2012-01-01

Background Striga species are noxious root hemi-parasitic weeds that debilitate cereal production in sub-Saharan Africa (SSA). Control options for Striga are limited and developing Striga resistant crop germplasm is regarded as the best and most sustainable control measure. Efforts to improve germplasm for Striga resistance by a non-Genetic Modification (GM) approach, for example by exploiting natural resistance, or by a GM approach are constrained by limited information on the biological processes underpinning host-parasite associations. Additionaly, a GM approach is stymied by lack of availability of candidate resistance genes for introduction into hosts and robust transformation methods to validate gene functions. Indeed, a majority of Striga hosts, the world’s most cultivated cereals, are recalcitrant to genetic transformation. In maize, the existing protocols for transformation and regeneration are tedious, lengthy, and highly genotype-specific with low efficiency of transformation. Results We used Agrobacterium rhizogenes strain K599 carrying a reporter gene construct, Green Fluorescent Protein (GFP), to generate transgenic composite maize plants that were challenged with the parasitic plant Striga hermonthica. Eighty five percent of maize plants produced transgenic hairy roots expressing GFP. Consistent with most hairy roots produced in other species, transformed maize roots exhibited a hairy root phenotype, the hallmark of A. rhizogenes mediated transformation. Transgenic hairy roots resulting from A. rhizogenes transformation were readily infected by S. hermonthica. There were no significant differences in the number and size of S. hermonthica individuals recovered from either transgenic or wild type roots. Conclusions This rapid, high throughput, transformation technique will advance our understanding of gene function in parasitic plant-host interactions. PMID:22720750

Enthalpic and Entropic Competition in Blends of Self-Suspended Hairy Nanoparticles

NASA Astrophysics Data System (ADS)

Choudhury, Snehashis; Agrawal, Akanksha; Archer, Lynden

Self-suspended hairy nanoparticles, where polymer chains are grafted onto nanoparticles, have attracted significant recent attention. These materials have been reported to manifest several interesting phenomena like thermal jamming, slowing-down of polymer chain dynamics, as well as small-strain stress overshoots during start-up of steady shear. The entropic penalty on tethered polymers produced by the requirement that they fill the space between the nanoparticle cores explain most of these behaviors. Here, we show that the entropic attraction between tethered polymer chains can be manipulated in mixtures of hairy nanoparticles using different polymer chemistry to design materials with unusual characteristics. Specifically, the degree of interpenetration of polymer chains can be controlled by tuning their interaction parameter (χ) . For SiO2-PEG/SiO2-PMMA blends, oscillatory rheological measurements show that the plateau modulus and yielding energy are significantly increased, while an opposite effect is seen with SiO2-PEG/SiO2-PI blends. More subtle effects of this enthalpy-entropy competition are well captured in Dielectric Spectroscopy measurements and SAXS experiments that can be used to quantify the degree of stretch and interdigitation of polymer chains.

9-methoxycanthin-6-one production in elicited hairy roots culture of Eurycoma longifolia

NASA Astrophysics Data System (ADS)

Abdullah, Nazirah; Ismail, Ismanizan; Hassan, Nor Hasnida; Basherudin, Norlia

2016-11-01

Eurycoma longifolia (Tongkat Ali) is a highly sought after medicinal plant in Malaysia. Propagation of E. longifolia through tissue culture has been reported in order to cater the industry demands for planting and raw materials as well as for conservation purposes. E. longifolia hairy roots culture has been developed using Agrobacterium rhizogenes for the production of Tongkat Ali phytochemicals. Effects of three elicitors; methyl jasmonate, salicylic acid, and yeast extract at different concentrations were evaluated on the production of 9-methoxycanthin-6-one in E. longifolia hairy roots. The cultures were elicited at early exponential growth phase, followed by extraction of 9-methoxycanthin-6-one using methanol and HPLC analysis. Elicitation with methyl jasmonate at all concentrations increased 9-methoxycanthin-6-one up to 1-3 fold and treatment with (0.1 mM) was most efficient in enhancing 9-methoxycanthin-6-one production up to 3.902 mg/g dry weight after 7 days (168 hours) elicitation.

Protein-functionalized hairy diamond nanoparticles.

PubMed

Dahoumane, Si Amar; Nguyen, Minh Ngoc; Thorel, Alain; Boudou, Jean-Paul; Chehimi, Mohamed M; Mangeney, Claire

2009-09-01

Diazonium salt chemistry and atom transfer radical polymerization (ATRP) were combined in view of preparing new bioactive hairy diamond nanoparticles containing, or potentially containing, nitrogen-vacancy (NV) fluorescent centers (fluorescent nanodiamonds, or fNDs). fNDs were modified by ATRP initiators using the electroless reduction of the diazonium salt BF(4)(-),(+)N(2)-C(6)H(4)-CH(CH(3))-Br. The strongly bound aryl groups -C(6)H(4)-CH(CH(3))-Br efficiently initiated the ATRP of tert-butyl methacrylate (tBMA) at the surface of the nanodiamonds, which resulted in obtaining ND-PtBMA hybrids. The grafted chain thickness, estimated from X-ray photoelectron spectroscopy (XPS), was found to increase linearly with respect to time before reaching a plateau value of ca. 2 nm. These nanoobjects were further hydrolyzed into ND-PMAA (where PMAA is the poly(methacrylic acid) graft) and further decorated by bovine serum albumin through the 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) coupling procedure.

Effectiveness of herbicides for control of hairy vetch (Vicia villosa) in winter wheat

USDA-ARS?s Scientific Manuscript database

We conducted a field experiment in 2009-10 at Pennsylvania and Maryland locations, and repeated it in 2010-11, to test the effectiveness of post-emergent herbicides applied at fall and spring timings on seeded hairy vetch in winter wheat. We tested 16 herbicide treatment combinations that included ...

Thyroid Hormone-Induced Activation of Notch Signaling is Required for Adult Intestinal Stem Cell Development During Xenopus Laevis Metamorphosis.

PubMed

Hasebe, Takashi; Fujimoto, Kenta; Kajita, Mitsuko; Fu, Liezhen; Shi, Yun-Bo; Ishizuya-Oka, Atsuko

2017-04-01

In Xenopus laevis intestine during metamorphosis, the larval epithelial cells are removed by apoptosis, and the adult epithelial stem (AE) cells appear concomitantly. They proliferate and differentiate to form the adult epithelium (Ep). Thyroid hormone (TH) is well established to trigger this remodeling by regulating the expression of various genes including Notch receptor. To study the role of Notch signaling, we have analyzed the expression of its components, including the ligands (DLL and Jag), receptor (Notch), and targets (Hairy), in the metamorphosing intestine by real-time reverse transcription-polymerase chain reaction and in situ hybridization or immunohistochemistry. We show that they are up-regulated during both natural and TH-induced metamorphosis in a tissue-specific manner. Particularly, Hairy1 is specifically expressed in the AE cells. Moreover, up-regulation of Hairy1 and Hairy2b by TH was prevented by treating tadpoles with a γ-secretase inhibitor (GSI), which inhibits Notch signaling. More importantly, TH-induced up-regulation of LGR5, an adult intestinal stem cell marker, was suppressed by GSI treatment. Our results suggest that Notch signaling plays a role in stem cell development by regulating the expression of Hairy genes during intestinal remodeling. Furthermore, we show with organ culture experiments that prolonged exposure of tadpole intestine to TH plus GSI leads to hyperplasia of secretory cells and reduction of absorptive cells. Our findings here thus provide evidence for evolutionarily conserved role of Notch signaling in intestinal cell fate determination but more importantly reveal, for the first time, an important role of Notch pathway in the formation of adult intestinal stem cells during vertebrate development. Stem Cells 2017;35:1028-1039. © 2016 The Authors STEM CELLS published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Increased synthesis of a new oleanane-type saponin in hairy roots of marigold (Calendula officinalis) after treatment with jasmonic acid.

PubMed

Markowski, Michał; Długosz, Marek; Szakiel, Anna; Durli, Mathieu; Poinsignon, Sophie; Bouguet-Bonnet, Sabine; Vernex-Loset, Lionel; Krier, Gabriel; Henry, Max

2018-04-18

Native plant of marigold (Calendula officinalis L.) synthesizes oleanolic acid saponins classified as glucosides or glucuronides according to the first residue in sugar chain bound to C-3 hydroxyl group. Hairy root culture, obtained by transformation with Agrobacterium rhizogenes strain 15834, exhibit a potent ability of synthesis of oleanolic acid glycosides. The HPLC profile of saponin fraction obtained from C. officinalis hairy roots treated with plant stress hormone, jasmonic acid, showed the 10-times increase of the content of one particular compound, determined by NMR and MALDI TOF as a new bisdesmoside saponin, 3-O-β-d-glucuronopyranosyl-28-O-β-d-galactopyranosyl-oleanolic acid. Such a diglycoside does not occur in native C. officinalis plant. It is a glucuronide, whereas in the native plant glucuronides are mainly accumulated in flowers, while glucosides are the most abundant saponins in roots. Thus, our results revealed that the pathways of saponin biosynthesis, particularly reactions of glycosylation, are altered in C. officinalis hairy root culture.

Enhanced Stilbene Production and Excretion in Vitis vinifera cv Pinot Noir Hairy Root Cultures.

PubMed

Tisserant, Leo-Paul; Aziz, Aziz; Jullian, Nathalie; Jeandet, Philippe; Clément, Christophe; Courot, Eric; Boitel-Conti, Michèle

2016-12-10

Stilbenes are defense molecules produced by grapevine in response to stresses including various elicitors and signal molecules. Together with their prominent role in planta, stilbenes have been the center of much attention in recent decades due to their pharmaceutical properties. With the aim of setting up a cost-effective and high purity production of resveratrol derivatives, hairy root lines were established from Vitis vinifera cv Pinot Noir 40024 to study the organ-specific production of various stilbenes. Biomass increase and stilbene production by roots were monitored during flask experiments. Although there was a constitutive production of stilbenes in roots, an induction of stilbene synthesis by methyl jasmonate (MeJA) after 18 days of growth led to further accumulation of ε-viniferin, δ-viniferin, resveratrol and piceid. The use of 100 µM MeJA after 18 days of culture in the presence of methyl-β-cyclodextrins (MCDs) improved production levels, which reached 1034µg/g fresh weight (FW) in roots and 165 mg/L in the extracellular medium, corresponding to five-and 570-foldincrease in comparison to control. Whereas a low level of stilbene excretion was measured in controls, addition of MeJA induced excretion of up to 37% of total stilbenes. The use of MCDs increased the excretion phenomenon even more, reaching up to 98%. Our results demonstrate the ability of grapevine hairy roots to produce various stilbenes. This production was significantly improved in response to elicitation by methyl jasmonate and/or MCDs. This supports the interest of using hairy roots as a potentially valuable system for producing resveratrol derivatives.

Scale-Up of Agrobacterium rhizogenes-Mediated Hairy Root Cultures of Rauwolfia serpentina: A Persuasive Approach for Stable Reserpine Production.

PubMed

Mehrotra, Shakti; Srivastava, Vikas; Goel, Manoj K; Kukreja, Arun K

2016-01-01

Roots of Rauwolfia serpentina, also known as "Sarpagandha" possess high pharmaceutical value due to the presence of reserpine and other medicinally important terpene indole alkaloids. Ever increasing commercial demand of R. serpentina roots is the major reason behind the unsystematic harvesting and fast decline of the species from its natural environment. Considering Agrobacterium rhizogenes-mediated hairy root cultures as an alternative source for the production of plant-based secondary metabolites, the present optimized protocol offers a commercially feasible method for the production of reserpine, the most potent alkaloid from R. serpentina roots. This end-to-end protocol presents the establishment of hairy root culture from the leaf explants of R. serpentina through the infection of A. rhizogenes strain A4 in liquid B5 culture medium and its up-scaling in a 5 L bench top, mechanically agitated bioreactor. The transformed nature of roots was confirmed through PCR-based rol A gene amplification in genomic DNA of putative hairy roots. The extraction and quantification of reserpine in bioreactor grown roots has been done using monolithic reverse phase high-performance liquid chromatography (HPLC).

Phytochelatin homologs induced in hairy roots of horseradish.

PubMed

Kubota, H; Sato, K; Yamada, T; Maitani, T

2000-01-01

When exposed to excess heavy metals, plants induce phytochelatins and related peptides (all designated as PCAs). Thus, when hairy roots of horseradish (Armoracia rusticana) were exposed for 3 days to cadmium (1 mM) along with reduced glutathione (2 mM), PCA induction occurred. Moreover, a new family of thiol peptides was detected as well as the previously known PCAs, as revealed by postcolumn-derivatization HPLC. Two were isolated and their structures were identified as (gamma-Glu-Cys)n-Gln (n = 3 and 4) by electrospray ionization-mass spectrometer spectra, this being confirmed by chemical synthesis of the peptides. These new analogs constitute the sixth PCA family identified to date.

Antibacterial, Anti-Inflammatory, Antioxidant, and Antiproliferative Properties of Essential Oils from Hairy and Normal Roots of Leonurus sibiricus L. and Their Chemical Composition.

PubMed

Sitarek, Przemysław; Rijo, Patricia; Garcia, Catarina; Skała, Ewa; Kalemba, Danuta; Białas, Adam J; Szemraj, Janusz; Pytel, Dariusz; Toma, Monika; Wysokińska, Halina; Śliwiński, Tomasz

2017-01-01

Essential oils obtained from the NR (normal roots) and HR (hairy roots) of the medicinal plant Leonurus sibiricus root were used in this study. The essential oil compositions were detected by GC-MS. Eighty-five components were identified in total. Seventy components were identified for NR essential oil. The major constituents in NR essential oil were β -selinene (9.9%), selina-4,7-diene (9.7%), (E) - β -caryophyllene (7.3%),myli-4(15)-ene (6.4%), and guaia-1(10),11-diene (5.9%). Sixty-seven components were identified in HR essential oil, the main constituents being (E) - β -caryophyllene (22.6%), and germacrene D (19.8%). The essential oils were tested for cytotoxic effect, antimicrobial, anti-inflammatory, and antioxidant activities. Both essential oils showed activity against grade IV glioma cell lines (IC 50 = 400  μ g/mL), antimicrobial (MIC and MFC values of 2500 to 125  μ g/mL), and anti-inflammatory (decreased level of IL-1 β , IL-6, TNF- α , and IFN- γ in LPS-stimulated cells).The essential oils exhibited moderate antioxidant activity in ABTS (EC 50 = 98 and 88  μ g/mL) assay. This is the first study to examine composition of the essential oils and their antimicrobial, antioxidant, antiproliferative, and anti-inflammatory activities. The results indicate that essential oils form L. sibiricus root may be used in future as an alternative to synthetic antimicrobial agents with potential application in the food and pharmaceutical industries.

Genetic diversity of resident soil rhizobia isolated from nodules of distinct hairy vetch genotypes

USDA-ARS?s Scientific Manuscript database

Hairy vetch (Vicia villosa Roth) is widely grown as a legume cover crop throughout the U.S.A., with biological nitrogen fixation (BNF) through symbiosis with Rhizobium leguminosarum biovar viciae (Rlv) being one of the most sought after benefits of its cultivation. This study determined if HV culti...

Hairy AdS black holes with a toroidal horizon in 4D Einstein-nonlinear σ-model system

NASA Astrophysics Data System (ADS)

Astorino, Marco; Canfora, Fabrizio; Giacomini, Alex; Ortaggio, Marcello

2018-01-01

An exact hairy asymptotically locally AdS black hole solution with a flat horizon in the Einstein-nonlinear sigma model system in (3+1) dimensions is constructed. The ansatz for the nonlinear SU (2) field is regular everywhere and depends explicitly on Killing coordinates, but in such a way that its energy-momentum tensor is compatible with a metric with Killing fields. The solution is characterized by a discrete parameter which has neither topological nor Noether charge associated with it and therefore represents a hair. A U (1) gauge field interacting with Einstein gravity can also be included. The thermodynamics is analyzed. Interestingly, the hairy black hole is always thermodynamically favoured with respect to the corresponding black hole with vanishing Pionic field.

Use of Model-Based Nutrient Feeding for Improved Production of Artemisinin by Hairy Roots of Artemisia Annua in a Modified Stirred Tank Bioreactor.

PubMed

Patra, Nivedita; Srivastava, Ashok K

2015-09-01

Artemisinin has been indicated to be a potent drug for the cure of malaria. Batch growth and artemisinin production kinetics of hairy root cultures of Artemisia annua were studied under shake flask conditions which resulted in accumulation of 12.49 g/L biomass and 0.27 mg/g artemisinin. Using the kinetic data, a mathematical model was identified to understand and optimize the system behavior. The developed model was then extrapolated to design nutrient feeding strategies during fed-batch cultivation for enhanced production of artemisinin. In one of the fed-batch cultivation, sucrose (37 g/L) feeding was done at a constant feed rate of 0.1 L/day during 10-15 days, which led to improved artemisinin accumulation of 0.77 mg/g. The second strategy of fed-batch hairy root cultivation involved maintenance of pseudo-steady state sucrose concentration (20.8 g/L) during 10-15 days which resulted in artemisinin accumulation of 0.99 mg/g. Fed-batch cultivation (with the maintenance of pseudo-steady state of substrate) of Artemisia annua hairy roots was, thereafter, implemented in bioreactor cultivation, which featured artemisinin accumulation of 1.0 mg/g artemisinin in 16 days of cultivation. This is the highest reported artemisinin yield by hairy root cultivation in a bioreactor.

Transgenic studies reveal the positive role of LeEIL-1 in regulating shikonin biosynthesis in Lithospermum erythrorhizon hairy roots.

PubMed

Fang, Rongjun; Zou, Ailan; Zhao, Hua; Wu, Fengyao; Zhu, Yu; Zhao, Hu; Liao, Yonghui; Tang, Ren-Jie; Pang, Yanjun; Yang, Rongwu; Wang, Xiaoming; Qi, Jinliang; Lu, Guihua; Yang, Yonghua

2016-05-26

The phytohormone ethylene (ET) is a key signaling molecule for inducing the biosynthesis of shikonin and its derivatives, which are secondary metabolites in Lithospermum erythrorhizon. Although ETHYLENE INSENSITIVE3 (EIN3)/EIN3-like proteins (EILs) are crucial transcription factors in ET signal transduction pathway, the possible function of EIN3/EIL1 in shikonin biosynthesis remains unknown. In this study, by targeting LeEIL-1 (L. erythrorhizon EIN3-like protein gene 1) at the expression level, we revealed the positive regulatory effect of LeEIL-1 on shikonin formation. The mRNA level of LeEIL-1 was significantly up-regulated and down-regulated in the LeEIL-1-overexpressing hairy root lines and LeEIL-1-RNAi hairy root lines, respectively. Specifically, LeEIL-1 overexpression resulted in increased transcript levels of the downstream gene of ET signal transduction pathway (LeERF-1) and a subset of genes for shikonin formation, excretion and/or transportation (LePAL, LeC4H-2, Le4CL-1, HMGR, LePGT-1, LeDI-2, and LePS-2), which was consistent with the enhanced shikonin contents in the LeEIL-1-overexpressing hairy root lines. Conversely, LeEIL-1-RNAi dramatically repressed the expression of the above genes and significantly reduced shikonin production. The results revealed that LeEIL-1 is a positive regulator of the biosynthesis of shikonin and its derivatives in L. erythrorhizon hairy roots. Our findings gave new insights into the molecular regulatory mechanism of ET in shikonin biosynthesis. LeEIL-1 could be a crucial target gene for the genetic engineering of shikonin biosynthesis.

Analysis of tomato gene promoters activated in syncytia induced in tomato and potato hairy roots by Globodera rostochiensis.

PubMed

Wiśniewska, A; Dąbrowska-Bronk, J; Szafrański, K; Fudali, S; Święcicka, M; Czarny, M; Wilkowska, A; Morgiewicz, K; Matusiak, J; Sobczak, M; Filipecki, M

2013-06-01

The potato cyst nematode (Globodera rostochiensis) induces feeding sites (syncytia) in tomato and potato roots. In a previous study, 135 tomato genes up-regulated during G. rostochiensis migration and syncytium development were identified. Five genes (CYP97A29, DFR, FLS, NIK and PMEI) were chosen for further study to examine their roles in plant-nematode interactions. The promoters of these genes were isolated and potential cis regulatory elements in their sequences were characterized using bioinformatics tools. Promoter fusions with the β-glucuronidase gene were constructed and introduced into tomato and potato genomes via transformation with Agrobacterium rhizogenes to produce hairy roots. The analysed promoters displayed different activity patterns in nematode-infected and uninfected transgenic hairy roots.

Hairy Lovelock black holes and Stueckelberg mechanism for Weyl symmetry

NASA Astrophysics Data System (ADS)

Chernicoff, Mariano; Giribet, Gaston; Oliva, Julio

2016-10-01

Lovelock theory of gravity -and, in particular, Einstein theory- admits black hole solutions that can be equipped with a hair by conformally coupling the theory to a real scalar field. This is a secondary hair, meaning that it does not endow the black hole with new quantum numbers. It rather consists of a non-trivial scalar field profile of fixed intensity which turns out to be regular everywhere outside and on the horizon and, provided the cosmological constant is negative, behaves at large distance in a way compatible with the Anti-de Sitter (AdS) asymptotic. In this paper, we review the main features of these hairy black hole solutions, such as their geometrical and thermodynamical properties. The conformal coupling to matter in dimension D > 4 in principle includes higher-curvature terms. These couplings are obtained from the Lovelock action through the Stueckelberg strategy. As a consequence, the resulting scalar-tensor theory exhibits a self-duality under field redefinition that resembles T-duality. Through this field redefinition, the matter content of the theory transforms into a Lovelock action for a dual geometry. Since the hairy black holes only exist for special relations between the dual Lovelock coupling constants, it is natural to compare those relations with the causality bounds coming from AdS/CFT. We observe that, while the lower causality bound is always obeyed, the upper causality bound is violated. The latter, however, is saturated in the large D limit.

Retrospective Evaluation of Hairy Cell Leukemia Patients Treated with Three Different First-Line Treatment Modalities in the Last Two Decades: A Single-Center Experience.

PubMed

Öngören, Şeniz; Eşkazan, Ahmet Emre; Berk, Selin; Elverdi, Tuğrul; Salihoğlu, Ayşe; Ar, Muhlis Cem; Başlar, Zafer; Aydın, Yıldız; Tüzüner, Nükhet; Soysal, Teoman

2017-12-01

In this study, we retrospectively analyzed the clinical outcome, treatment responses, infectious complications, and survival rates of 71 hairy cell leukemia (HCL) cases. Sixty-seven patients received a first-line treatment and 2-chlorodeoxyadenosine (cladribine-2-CdA) was administered in 31 cases, 19 patients received interferon-alpha (INF-α), splenectomy was performed in 16 cases, and rituximab was used in one. Although the highest overall response rate (ORR) was observed in patients receiving 2-CdA upfront, ORRs were comparable in the 2-CdA, INF-α, and splenectomy subgroups. Relapse rates were significantly lower in patients who received first-line 2-CdA. The progression-free survival (PFS) rate with 2-CdA was significantly higher than in patients with INF-α and splenectomy, but we found similar overall survival rates with all three upfront treatment modalities. Infections including tuberculosis were a major problem. Although purine analogues have improved the ORRs and PFS, there is still much progress to make with regard to overall survival and relapsed/refractory disease in patients with HCL.

Estrogen-dependent downregulation of hairy and enhancer of split homolog-1 gene expression in breast cancer cells is mediated via a 3' distal element.

PubMed

Müller, Patrick; Merrell, Kenneth W; Crofts, Justin D; Rönnlund, Caroline; Lin, Chin-Yo; Gustafsson, Jan-Ake; Ström, Anders

2009-03-01

Regulation of hairy and enhancer of split homologue-1 (HES-1) by estradiol and all-trans retinoic acid affects proliferation of human breast cancer cells. Here, we identify and characterize cis-regulatory elements involved in HES-1 regulation. In the distal 5' promoter of the HES-1 gene, we found a retinoic acid response element and in the distal 3' region, an estrogen receptor alpha(ER)alpha binding site. The ERalpha binding site, composed of an estrogen response element (ERE) and an ERE half-site, is important for both ERalpha binding and transcriptional regulation. Chromatin immunoprecipitation assays revealed that ERalpha is recruited to the ERE and associates with the HES-1 promoter. We also show recruitment of nuclear receptor co-regulators to the ERE in response to estradiol, followed by a decrease in histone acetylation and RNA polymerase II docking in the HES-1 promoter region. Our findings are consistent with a novel type of repressive estrogen response element in the distal 3' region of the HES-1 gene.

Enhanced triterpene accumulation in Panax ginseng hairy roots overexpressing mevalonate-5-pyrophosphate decarboxylase and farnesyl pyrophosphate synthase.

PubMed

Kim, Yong-Kyoung; Kim, Yeon Bok; Uddin, Md Romij; Lee, Sanghyun; Kim, Soo-Un; Park, Sang Un

2014-10-17

To elucidate the function of mevalonate-5-pyrophosphate decarboxylase (MVD) and farnesyl pyrophosphate synthase (FPS) in triterpene biosynthesis, the genes governing the expression of these enzymes were transformed into Panax ginseng hairy roots. All the transgenic lines showed higher expression levels of PgMVD and PgFPS than that by the wild-type control. Among the hairy root lines transformed with PgMVD, M18 showed the highest level of transcription compared to the control (14.5-fold higher). Transcriptions of F11 and F20 transformed with PgFPS showed 11.1-fold higher level compared with control. In triterpene analysis, M25 of PgMVD produced 4.4-fold higher stigmasterol content (138.95 μg/100 mg, dry weight [DW]) than that by the control; F17 of PgFPS showed the highest total ginsenoside (36.42 mg/g DW) content, which was 2.4-fold higher compared with control. Our results indicate that metabolic engineering in P. ginseng was successfully achieved through Agrobacterium rhizogenes-mediated transformation and that the accumulation of phytosterols and ginsenosides was enhanced by introducing the PgMVD and PgFPS genes into the hairy roots of the plant. Our results suggest that PgMVD and PgFPS play an important role in the triterpene biosynthesis of P. ginseng.

[Research of mechanism of secondary metabolites of phenolic acids in Salvia miltiorrhiza hairy root induced by jasmonate].

PubMed

Li, Wenyuan; Gao, Wei; Zhao, Jing; Cui, Guanghong; Shao, Aijuan; Huang, Luqi

2012-01-01

To study the mechanism of secondary metabolites of some phenolic acids in the hairy roots of Salvia miltiorrhiza induced by methyl jasmonate. The hairy roots of S. miltiorrhiza were induced with methyl jasmonate (100 micromol x L(-1)) and collected at 0, 12, 24, 36 h after treatment. Real-time quantitative PCR was used for detecting the mRNA expression level of the key enzyme genes on the secondary metabolites pathway of rosmarinic acid, while a LC-MS method was developed to determine the content of rosmarinic acid, caffeic acid and salvianolic acid B. The concentration of phenolic acids grew up and accumulated quickly in the hairy roots with exogenous signal molecule MJ induced, and it was showed that the content of CA and RA reached the maximum after 24 h and the content of LAB reached the maximum in 36 h by MJ induced. The induction mechanism may be activated with different levels of RA synthesis in PAL, 4CL, C4H genes on the key enzyme phenylalanine pathway and TAT, HPPR genes on tyrosine pathway. The time of gene expression was different, among them, 4CL and PAL genes were more important. In a word, the result can provide some basis data about the mechanism of secondary metabolites of phenolic acids for further research.

Skin-derived dendritic cells acquire and degrade the scrapie agent following in vitro exposure

PubMed Central

Mohan, Joanne; Hopkins, John; Mabbott, Neil A

2005-01-01

The accumulation of the scrapie agent in lymphoid tissues following inoculation via the skin is critical for efficient neuroinvasion, but how the agent is initially transported from the skin to the draining lymph node is not known. Langerhans cells (LCs) are specialized antigen-presenting cells that continually sample their microenvironment within the epidermis and transport captured antigens to draining lymph nodes. We considered LCs probable candidates to acquire and transport the scrapie agent after inoculation via the skin. XS106 cells are dendritic cells (DCs) isolated from mouse epidermis with characteristics of mature LC cells. To investigate the potential interaction of LCs with the scrapie agent XS106 cells were exposed to the scrapie agent in vitro. We show that XS106 cells rapidly acquire the scrapie agent following in vitro exposure. In addition, XS106 cells partially degrade the scrapie agent following extended cultivation. These data suggest that LCs might acquire and degrade the scrapie agent after inoculation via the skin, but data from additional experiments demonstrate that this ability could be lost in the presence of lipopolysaccharide or other immunostimulatory molecules. Our studies also imply that LCs would not undergo maturation following uptake of the scrapie agent in the skin, as the expression of surface antigens associated with LC maturation were unaltered following exposure. In conclusion, although LCs or DCs have the potential to acquire the scrapie agent within the epidermis our data suggest it is unlikely that they become activated and stimulated to transport the agent to the draining lymph node. PMID:16108824

Ablation of human skin mast cells in situ by lysosomotropic agents.

PubMed

Hagforsen, Eva; Paivandy, Aida; Lampinen, Maria; Weström, Simone; Calounova, Gabriela; Melo, Fabio R; Rollman, Ola; Pejler, Gunnar

2015-07-01

Mast cells are known to have a detrimental impact on numerous types of inflammatory skin diseases such as contact dermatitis, atopic eczema and cutaneous mastocytosis. Regimens that dampen skin mast cell-mediated activities can thus offer an attractive therapeutic option under such circumstances. As mast cells are known to secrete a large array of potentially pathogenic compounds, both from preformed stores in secretory lysosomes (granules) and after de novo synthesis, mere inhibition of degranulation or interference with individual mast cell mediators may not be sufficient to provide an effective blockade of harmful mast cell activities. An alternative strategy may therefore be to locally reduce skin mast cell numbers. Here, we explored the possibility of using lysosomotropic agents for this purpose, appreciating the fact that mast cell granules contain bioactive compounds prone to trigger apoptosis if released into the cytosolic compartment. Based on this principle, we show that incubation of human skin punch biopsies with the lysosomotropic agents siramesine or Leu-Leu methyl ester preferably ablated the mast cell population, without causing any gross adverse effects on the skin morphology. Subsequent analysis revealed that mast cells treated with lysosomotropic agents predominantly underwent apoptotic rather than necrotic cell death. In summary, this study raises the possibility of using lysosomotropic agents as a novel approach to targeting deleterious mast cell populations in cutaneous mastocytosis and other skin disorders negatively influenced by mast cells. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Biomass Production of Hairy Roots of Artemisia annua and Arachis hypogaea in a Scaled-Up Mist Bioreactor

PubMed Central

Sivakumar, Ganapathy; Liu, Chunzhao; Towler, Melissa J.

2014-01-01

Hairy roots have the potential to produce a variety of valuable small and large molecules. The mist reactor is a gas phase bioreactor that has shown promise for low-cost culture of hairy roots. Using a newer, disposable culture bag, mist reactor performance was studied with two species, Artemisia annua L. and Arachis hypogaea (peanut), at scales from 1 to 20 L. Both species of hairy roots when grown at 1 L in the mist reactor showed growth rates that surpassed that in shake flasks. From the information gleaned at 1 L, Arachis was scaled further to 4 and then 20 L. Misting duty cycle, culture medium flow rate, and timing of when flow rate was increased were varied. In a mist reactor increasing the misting cycle or increasing the medium flow rate are the two alternatives for increased delivery of liquid nutrients to the root bed. Longer misting cycles beyond 2–3 min were generally deemed detrimental to growth. On the other hand, increasing the medium flow rate to the sonic nozzle especially during the exponential phase of root growth (weeks 2–3) was the most important factor for increasing growth rates and biomass yields in the 20 L reactors. A. hypogaea growth in 1 L reactors was μ = 0.173 day−1 with biomass yield of 12.75 g DWL−1. This exceeded that in shake flasks at μ = 0.166 day−1 and 11.10 g DWL−1. Best growth rate and biomass yield at 20 L was μ = 0.147 and 7.77 g DWL−1, which was mainly achieved when medium flow rate delivery was increased. The mist deposition model was further evaluated using this newer reactor design and when the apparent thickness of roots (+hairs) was taken into account, the empirical data correlated with model predictions. Together these results establish the most important conditions to explore for future optimization of the mist bioreactor for culture of hairy roots. PMID:20687140

Modification of phenolic metabolism in soybean hairy roots through down regulation of chalcone synthase or isoflavone synthase.

PubMed

Lozovaya, Vera V; Lygin, Anatoliy V; Zernova, Olga V; Ulanov, Alexander V; Li, Shuxian; Hartman, Glen L; Widholm, Jack M

2007-02-01

Soybean hairy roots, transformed with the soybean chalcone synthase (CHS6) or isoflavone synthase (IFS2) genes, with dramatically decreased capacity to synthesize isoflavones were produced to determine what effects these changes would have on susceptibility to a fungal pathogen. The isoflavone and coumestrol concentrations were decreased by about 90% in most lines apparently due to gene silencing. The IFS2 transformed lines had very low IFS enzyme activity in microsomal fractions as measured by the conversion of naringenin to genistein. The CHS6 lines with decreased isoflavone concentrations had 5 to 20-fold lower CHS enzyme activities than the appropriate controls. Both IFS2 and CHS transformed lines accumulated higher concentrations of both soluble and cell wall bound phenolic acids compared to controls with higher levels found in the CHS6 lines indicating alterations in the lignin biosynthetic branch of the pathway. Induction of the soybean phytoalexin glyceollin, of which the precursor is the isoflavone daidzein, by the fungal pathogen Fusarium solani f. sp. glycines (FSG) that causes soybean sudden death syndrome (SDS) showed that the low isoflavone transformed lines did not accumulate glyceollin while the control lines did. The (iso)liquritigenin content increased upon FSG induction in the IFS2 transformed roots indicating that the pathway reactions before this point can control isoflavonoid synthesis. The lowest fungal growth rate on hairy roots was found on the FSG partially resistant control roots followed by the SDS sensitive control roots and the low isoflavone transformants. The results indicate the importance of phytoalexin synthesis in root resistance to the pathogen.

16D10 siRNAs inhibit root-knot nematode infection in transgenic grape hairy roots

USDA-ARS?s Scientific Manuscript database

To develop a biotech-based solution for controlling Root-knot nematodes (RKNs) in grapes, we evaluated the efficacy of plant-derived RNA interference (RNAi) silencing of a conserved RKN effector gene, 16D10, for nematode resistance in transgenic grape hairy roots. Two hairpin-based silencing constru...

Effect of elicitors on the production of gossypol and methylated gossypol in cotton hairy roots

USDA-ARS?s Scientific Manuscript database

The effect of two-chemical elicitors, salicylic acid and methyl jasmonate, on the production of gossypol, 6-methoxy gossypol, and 6,6'-dimethoxy gossypol in Gossypium barbadense hairy roots was examined. Methyl jasmonate, but not salicylic acid, was found to increase the production of gossypol and ...

Labdane-type diterpenoids from hairy root cultures of Coleus forskohlii, possible intermediates in the biosynthesis of forskolin.

PubMed

Asada, Yoshihisa; Li, Wei; Terada, Tomohiro; Kuang, Xinzhu; Li, Qin; Yoshikawa, Takafumi; Hamaguchi, Shogo; Namekata, Iyuki; Tanaka, Hikaru; Koike, Kazuo

2012-07-01

Significant attention has been devoted to studying hairy root cultures as a promising strategy for production of various valuable secondary metabolites. These offer many advantages, such as high growth rate, genetic stability and being hormone-free. In this study, a detailed phytochemical investigation of the secondary metabolites of Coleus forskohlii hairy root cultures was undertaken and which resulted in the isolation of 22 compounds, including four forskolin derivatives and a monoterpene. Their structures were elucidated by extensive spectroscopic analyses. These compounds could be classified into four groups viz.: labdane-type diterpenes, monoterpenes, triterpenes and phenylpropanoid dimers. Apart from one compound, all labdane type diterpenes are oxygenated at C-11 as in forskolin and a scheme showing their biosynthetic relationships is proposed. Copyright © 2012 Elsevier Ltd. All rights reserved.

Agent-Based Computational Modeling of Cell Culture ...

EPA Pesticide Factsheets

Quantitative characterization of cellular dose in vitro is needed for alignment of doses in vitro and in vivo. We used the agent-based software, CompuCell3D (CC3D), to provide a stochastic description of cell growth in culture. The model was configured so that isolated cells assumed a “fried egg shape” but became increasingly cuboidal with increasing confluency. The surface area presented by each cell to the overlying medium varies from cell-to-cell and is a determinant of diffusional flux of toxicant from the medium into the cell. Thus, dose varies among cells for a given concentration of toxicant in the medium. Computer code describing diffusion of H2O2 from medium into each cell and clearance of H2O2 was calibrated against H2O2 time-course data (25, 50, or 75 uM H2O2 for 60 min) obtained with the Amplex Red assay for the medium and the H2O2-sensitive fluorescent reporter, HyPer, for cytosol. Cellular H2O2 concentrations peaked at about 5 min and were near baseline by 10 min. The model predicted a skewed distribution of surface areas, with between cell variation usually 2 fold or less. Predicted variability in cellular dose was in rough agreement with the variation in the HyPer data. These results are preliminary, as the model was not calibrated to the morphology of a specific cell type. Future work will involve morphology model calibration against human bronchial epithelial (BEAS-2B) cells. Our results show, however, the potential of agent-based modeling

Effect of light on the kinetics and equilibrium of the textile dye (Reactive Red 120) adsorption by Helianthus annuus hairy roots.

PubMed

Srikantan, Chitra; Suraishkumar, G K; Srivastava, Smita

2018-06-01

The study demonstrates for the first time that light influences the adsorption equilibrium and kinetics of a dye by root culture system. The azo dye (Reactive Red 120) adsorption by the hairy roots of H. annuus followed a pseudo first-order kinetic model and the adsorption equilibrium parameters were best estimated using Langmuir isotherm. The maximum dye adsorption capacity of the roots increased 6-fold, from 0.26 mg g -1 under complete dark conditions to 1.51 mg g -1 under 16/8 h light/dark photoperiod. Similarly, adsorption rate of the dye and removal (%) also increased in the presence of light, irrespective of the initial concentration of the dye (20-110 mg L -1 ). The degradation of the azo dye upon adsorption by the hairy roots of H. annuus was also confirmed. In addition, a strategy for simultaneous dye removal and increased alpha-tocopherol (industrially relevant) production by H. annuus hairy root cultures has been proposed and demonstrated. Copyright © 2018 Elsevier Ltd. All rights reserved.

New cancer cells apoptosis agents: Fluorinated aza-heterocycles

NASA Astrophysics Data System (ADS)

Prima, D. O.; Baev, D. S.; Vorontsova, E. V.; Frolova, T. S.; Bagryanskaya, I. Yu.; Slizhov, Yu. G.; Tolstikova, T. G.; Makarov, A. Yu.; Zibarev, A. V.

2017-09-01

Fluorinated benzo-fused 1,3-diazoles, 1,2,3-triazoles, 1,2,5-thia/selenadiazoles and 1,4-diazines were synthesized and tried for cytotoxicity towards the Hep2 (laryngeal epidermoid carcinoma) cells. The diazoles, triazoles and selenadiazoles were cytotoxic with IC50 = 2.2-26.4 µM and induced the cells apoptosis at concentrations C = 1-25 µM. At the same time, they were nontoxic towards normal cells. Due to this, these scaffolds were used in the computer-aided molecular design of new antitumor agents. Particularly, novel 1,2,3-triazole and 1,3-diazole derivatives for the binding site of the PAS domain of the transcription factor HIF were designed and some of them synthesized for further study. Overall, new anticancer agents featuring apoptotic activity are suggested.

Expression of a Recombinant Anti-HIV and Anti-Tumor Protein, MAP30, in Nicotiana tobacum Hairy Roots: A pH-Stable and Thermophilic Antimicrobial Protein.

PubMed

Moghadam, Ali; Niazi, Ali; Afsharifar, Alireza; Taghavi, Seyed Mohsen

2016-01-01

In contrast to conventional antibiotics, which microorganisms can readily evade, it is nearly impossible for a microbial strain that is sensitive to antimicrobial proteins to convert to a resistant strain. Therefore, antimicrobial proteins and peptides that are promising alternative candidates for the control of bacterial infections are under investigation. The MAP30 protein of Momordica charantia is a valuable type I ribosome-inactivating protein (RIP) with anti-HIV and anti-tumor activities. Whereas the antimicrobial activity of some type I RIPs has been confirmed, less attention has been paid to the antimicrobial activity of MAP30 produced in a stable, easily handled, and extremely cost-effective protein-expression system. rMAP30-KDEL was expressed in Nicotiana tobacum hairy roots, and its effect on different microorganisms was investigated. Analysis of the extracted total proteins of transgenic hairy roots showed that rMAP30-KDEL was expressed effectively and that this protein exhibited significant antibacterial activity in a dose-dependent manner. rMAP30-KDEL also possessed thermal and pH stability. Bioinformatic analysis of MAP30 and other RIPs regarding their conserved motifs, amino-acid contents, charge, aliphatic index, GRAVY value, and secondary structures demonstrated that these factors accounted for their thermophilicity. Therefore, RIPs such as MAP30 and its derived peptides might have promising applications as food preservatives, and their analysis might provide useful insights into designing clinically applicable antibiotic agents.

Expression of a Recombinant Anti-HIV and Anti-Tumor Protein, MAP30, in Nicotiana tobacum Hairy Roots: A pH-Stable and Thermophilic Antimicrobial Protein

PubMed Central

Moghadam, Ali; Niazi, Ali; Afsharifar, Alireza; Taghavi, Seyed Mohsen

2016-01-01

In contrast to conventional antibiotics, which microorganisms can readily evade, it is nearly impossible for a microbial strain that is sensitive to antimicrobial proteins to convert to a resistant strain. Therefore, antimicrobial proteins and peptides that are promising alternative candidates for the control of bacterial infections are under investigation. The MAP30 protein of Momordica charantia is a valuable type I ribosome-inactivating protein (RIP) with anti-HIV and anti-tumor activities. Whereas the antimicrobial activity of some type I RIPs has been confirmed, less attention has been paid to the antimicrobial activity of MAP30 produced in a stable, easily handled, and extremely cost-effective protein-expression system. rMAP30-KDEL was expressed in Nicotiana tobacum hairy roots, and its effect on different microorganisms was investigated. Analysis of the extracted total proteins of transgenic hairy roots showed that rMAP30-KDEL was expressed effectively and that this protein exhibited significant antibacterial activity in a dose-dependent manner. rMAP30-KDEL also possessed thermal and pH stability. Bioinformatic analysis of MAP30 and other RIPs regarding their conserved motifs, amino-acid contents, charge, aliphatic index, GRAVY value, and secondary structures demonstrated that these factors accounted for their thermophilicity. Therefore, RIPs such as MAP30 and its derived peptides might have promising applications as food preservatives, and their analysis might provide useful insights into designing clinically applicable antibiotic agents. PMID:27459300

Antibacterial, Anti-Inflammatory, Antioxidant, and Antiproliferative Properties of Essential Oils from Hairy and Normal Roots of Leonurus sibiricus L. and Their Chemical Composition

PubMed Central

Rijo, Patricia; Garcia, Catarina; Kalemba, Danuta; Szemraj, Janusz; Pytel, Dariusz; Toma, Monika; Śliwiński, Tomasz

2017-01-01

Essential oils obtained from the NR (normal roots) and HR (hairy roots) of the medicinal plant Leonurus sibiricus root were used in this study. The essential oil compositions were detected by GC-MS. Eighty-five components were identified in total. Seventy components were identified for NR essential oil. The major constituents in NR essential oil were β-selinene (9.9%), selina-4,7-diene (9.7%), (E)-β-caryophyllene (7.3%),myli-4(15)-ene (6.4%), and guaia-1(10),11-diene (5.9%). Sixty-seven components were identified in HR essential oil, the main constituents being (E)-β-caryophyllene (22.6%), and germacrene D (19.8%). The essential oils were tested for cytotoxic effect, antimicrobial, anti-inflammatory, and antioxidant activities. Both essential oils showed activity against grade IV glioma cell lines (IC50 = 400 μg/mL), antimicrobial (MIC and MFC values of 2500 to 125 μg/mL), and anti-inflammatory (decreased level of IL-1β, IL-6, TNF-α, and IFN-γ in LPS-stimulated cells).The essential oils exhibited moderate antioxidant activity in ABTS (EC50 = 98 and 88 μg/mL) assay. This is the first study to examine composition of the essential oils and their antimicrobial, antioxidant, antiproliferative, and anti-inflammatory activities. The results indicate that essential oils form L. sibiricus root may be used in future as an alternative to synthetic antimicrobial agents with potential application in the food and pharmaceutical industries. PMID:28191277

Agent-Based Modeling of Cancer Stem Cell Driven Solid Tumor Growth.

PubMed

Poleszczuk, Jan; Macklin, Paul; Enderling, Heiko

2016-01-01

Computational modeling of tumor growth has become an invaluable tool to simulate complex cell-cell interactions and emerging population-level dynamics. Agent-based models are commonly used to describe the behavior and interaction of individual cells in different environments. Behavioral rules can be informed and calibrated by in vitro assays, and emerging population-level dynamics may be validated with both in vitro and in vivo experiments. Here, we describe the design and implementation of a lattice-based agent-based model of cancer stem cell driven tumor growth.

Replication and spread of CJD, kuru and scrapie agents in vivo and in cell culture

PubMed Central

Miyazawa, Kohtaro; Emmerling, Kaitlin

2011-01-01

Transmissible Spongiform Encephalopathy (TSE) agents are defined by their virulence for particular species, their spread in the population, their incubation time to cause disease, and their neuropathological sequelae. Murine adapted human agents, including sporadic CJD (sCJD), New Guinea kuru, and Japanese CJD agents, display particularly distinct incubation times and maximal infectious brain titers. They also induce agent-specific patterns of neurodegeneration. When these TSE agents are transmitted to cultured hypothalamic GT1 cells they maintain their unique identities. Nevertheless, the human kuru (kCJD) and Japanese FU-CJD agents, as well as the sheep 22L and 263K scrapie agents display doubling times that are 8× to 33× faster in cells than in brain, indicating release from complex innate immune responses. These data are most consistent with a foreign viral structure, rather than an infectious form of host prion protein (PrP-res). Profound agent-specific inhibitory effects are also apparent in GT1 cells, and maximal titer plateau in kCJD and FU-CJD differed by 1,000-fold in a cell-based assay. Remarkably, the lower titer kCJD agent rapidly induced de novo PrP-res in GT1 cells, whereas the high titer FU-CJD agent replicated silently for multiple passages. Although PrP-res is often considered to be toxic, PrP-res instead may be part of a primal defense and/or clearance mechanism against TSE environmental agents. Limited spread of particular TSE agents through nanotubes and cell-to-cell contacts probably underlies the long peripheral phase of human CJD. PMID:21527829

Research on DNA methylation of human osteosarcoma cell MGMT and its relationship with cell resistance to alkylating agents.

PubMed

Guo, Jun; Cui, Qiu; Jiang, WeiHao; Liu, Cheng; Li, DingFeng; Zeng, Yanjun

2013-08-01

The objective of this study was to explore the O(6)-methylguanine-DNA methyltransferase (MGMT) gene methylation status and its protein expression, as well as the effects of demethylating agent 5-Aza-2'-deoxycytidine (5-Aza-CdR) on MGMT gene expression and its resistance to alkylating agents, and to elucidate MGMT expression mechanism and significance in osteosarcoma. The human osteosarcoma cell lines Saos-2 and MG-63 were collected and treated with 5-Aza-CdR for 6 days. The cells not treated with 5-Aza-CdR were set as a negative control. The genomic DNA was extracted from the Saos-2 and MG-63 cells using methylation-specific PCR to detect the promoter CpG island methylation status of the MGMT gene. Cell sensitivity to alkylating agents before and after drug administration was detected by the MTT method. The variation in MGMT gene mRNA and protein was detected by reverse transcription PCR (RT-PCR) and Western blotting. The MGMT promoter gene of normal Saos-2 cells was methylated, with reduced MGMT mRNA and protein expression; the MGMT mRNA and protein expression of Saos-2 cells treated with 5-Aza-CdR was obviously enhanced, and its sensitivity to alkylating agents was reversed. Meanwhile, with promoter CpG island unmethylation of the MGMT gene, MGMT protein was expressed in the normal MG-63 cells and the MG-63 cells treated with 5-Aza-CdR, and both showed resistance to alkylating agents. The methylation status of the MGMT gene promoter in human osteosarcoma cells reflected the cells' ability to induce MGMT protein expression and can be used as a molecular marker to project the sensitivity of cancer tissues to alkylating agent drugs.

Regulation of sesquiterpenoid metabolism in recombinant and elicited Valeriana officinalis hairy roots.

PubMed

Ricigliano, Vincent; Kumar, Santosh; Kinison, Scott; Brooks, Christopher; Nybo, S Eric; Chappell, Joe; Howarth, Dianella G

2016-05-01

The medicinal properties of Valerian (Valeriana officinalis) root preparations are attributed to the anxiolytic sesquiterpenoid valerenic acid and its biosynthetic precursors valerenal and valerenadiene, as well as the anti-inflammatory sesquiterpenoid β-caryophyllene. In order to study and engineer the biosynthesis of these pharmacologically active metabolites, a binary vector co-transformation system was developed for V. officinalis hairy roots. The relative expression levels and jasmonate-inducibility of a number of genes associated with sesquiterpenoid metabolism were profiled in roots: farnesyl pyrophosphate synthase (VoFPS), valerendiene synthase (VoVDS), germacrene C synthase (VoGCS), and a cytochrome P450 (CYP71D442) putatively associated with terpene metabolism based on sequence homology. Recombinant hairy root lines overexpressing VoFPS or VoVDS were generated and compared to control cultures. Overexpression of the VoFPS cDNA increased levels of the corresponding transcript 4- to 8-fold and sesquiterpene hydrocarbon accumulation by 1.5- to 4-fold. Overexpression of the VoVDS cDNA increased the corresponding transcript levels 5- to 9-fold and markedly increased yields of the oxygenated sesquiterpenoids valerenic acid and valerenal. Our findings suggest that the availability of cytoplasmic farnesyl diphosphate and valerenadiene are potential bottlenecks in Valeriana-specific sesquiterpenoid biosynthesis, which is also subject to regulation by methyl jasmonate elicitation. Copyright © 2016 Elsevier Ltd. All rights reserved.

Hairy black hole stability in AdS, quantum mechanics on the half-line and holography

NASA Astrophysics Data System (ADS)

Anabalón, Andrés; Astefanesei, Dumitru; Oliva, Julio

2015-10-01

We consider the linear stability of 4-dimensional hairy black holes with mixed boundary conditions in Anti-de Sitter spacetime. We focus on the mass of scalar fields around the maximally supersymmetric vacuum of the gauged N=8 supergravity in four dimensions, m 2 = -2 l -2. It is shown that the Schrödinger operator on the half-line, governing the S 2, H 2 or {{R}}^2 invariant mode around the hairy black hole, allows for non-trivial self-adjoint extensions and each of them corresponds to a class of mixed boundary conditions in the gravitational theory. Discarding the self-adjoint extensions with a negative mode impose a restriction on these boundary conditions. The restriction is given in terms of an integral of the potential in the Schrödinger operator resembling the estimate of Simon for Schrödinger operators on the real line. In the context of AdS/CFT duality, our result has a natural interpretation in terms of the field theory dual effective potential.

New alkaloids of the sarpagine group from Rauvolfia serpentina hairy root culture.

PubMed

Sheludko, Yuri; Gerasimenko, Irina; Kolshorn, Heinz; Stöckigt, Joachim

2002-07-01

Three new monoterpenoid indole alkaloids, 19(S),20(R)-dihydroperaksine (1), 19(S),20(R)-dihydroperaksine-17-al (2), and 10-hydroxy-19(S),20(R)-dihydroperaksine (3), along with 16 known alkaloids 4-19 were isolated from hairy root culture of Rauvolfia serpentina, and their structures were elucidated by 1D and 2D NMR analyses. Taking into account the stereochemistry of the new alkaloids and results of preliminary enzymatical studies, the putative biosynthetical relationships between the novel alkaloids are discussed.

Notch pathway signaling in the skin antagonizes Merkel cell development.

PubMed

Logan, Gregory J; Wright, Margaret C; Kubicki, Adam C; Maricich, Stephen M

2018-02-15

Merkel cells are mechanosensitive skin cells derived from the epidermal lineage whose development requires expression of the basic helix-loop-helix transcription factor Atoh1. The genes and pathways involved in regulating Merkel cell development during embryogenesis are poorly understood. Notch pathway signaling antagonizes Atoh1 expression in many developing body regions, so we hypothesized that Notch signaling might inhibit Merkel cell development. We found that conditional, constitutive overexpression of the Notch intracellular domain (NICD) in mouse epidermis significantly decreased Merkel cell numbers in whisker follicles and touch domes of hairy skin. Conversely, conditional deletion of the obligate NICD binding partner RBPj in the epidermis significantly increased Merkel cell numbers in whisker follicles, led to the development of ectopic Merkel cells outside of touch domes in hairy skin epidermis, and altered the distribution of Merkel cells in touch domes. Deletion of the downstream Notch effector gene Hes1 also significantly increased Merkel cell numbers in whisker follicles. Together, these data demonstrate that Notch signaling regulates Merkel cell production and patterning. Copyright © 2018 Elsevier Inc. All rights reserved.

Dynamic Nuclear Polarization NMR in Human Cells Using Fluorescent Polarizing Agents.

PubMed

Albert, Brice J; Gao, Chukun; Sesti, Erika L; Saliba, Edward P; Alaniva, Nicholas; Scott, Faith J; Sigurdsson, Snorri Th; Barnes, Alexander B

2018-06-20

Solid-state nuclear magnetic resonance (NMR) enables atomic resolution characterization of molecular structure and dynamics within complex heterogeneous samples, but it is typically insensitive. Dynamic nuclear polarization (DNP) increases NMR signal intensity by orders of magnitude and can be performed in combination with magic angle spinning (MAS) for sensitive, high-resolution spectroscopy. Here we report MAS DNP experiments, for the first time, within intact human cells with >40-fold DNP enhancement and a sample temperature below 6 K. In addition to cryogenic MAS results below 6 K, we also show in-cell DNP enhancements of 57-fold at 90 K. In-cell DNP is demonstrated using biradicals and sterically-shielded monoradicals as polarizing agents. A novel trimodal polarizing agent is introduced for DNP, which contains a nitroxide biradical, a targeting peptide for cell penetration, and a fluorophore for subcellular localization with confocal microscopy. The fluorescent polarizing agent provides in-cell DNP enhancements of 63-fold at a concentration of 2.7 mM. These experiments pave the way for structural characterization of biomolecules in an endogenous cellular context.

Enzymes and other agents that enhance cell wall extensibility

NASA Technical Reports Server (NTRS)

Cosgrove, D. J.

1999-01-01

Polysaccharides and proteins are secreted to the inner surface of the growing cell wall, where they assemble into a network that is mechanically strong, yet remains extensible until the cells cease growth. This review focuses on the agents that directly or indirectly enhance the extensibility properties of growing walls. The properties of expansins, endoglucanases, and xyloglucan transglycosylases are reviewed and their postulated roles in modulating wall extensibility are evaluated. A summary model for wall extension is presented, in which expansin is a primary agent of wall extension, whereas endoglucanases, xyloglucan endotransglycosylase, and other enzymes that alter wall structure act secondarily to modulate expansin action.

Suprafenacine, an Indazole-Hydrazide Agent, Targets Cancer Cells Through Microtubule Destabilization

PubMed Central

Choi, Bo-Hwa; Chattopadhaya, Souvik; Thanh, Le Nguyen; Feng, Lin; Nguyen, Quoc Toan; Lim, Chuan Bian; Harikishore, Amaravadhi; Nanga, Ravi Prakash Reddy; Bharatham, Nagakumar; Zhao, Yan; Liu, Xuewei; Yoon, Ho Sup

2014-01-01

Microtubules are a highly validated target in cancer therapy. However, the clinical development of tubulin binding agents (TBA) has been hampered by toxicity and chemoresistance issues and has necessitated the search for new TBAs. Here, we report the identification of a novel cell permeable, tubulin-destabilizing molecule - 4,5,6,7-tetrahydro-1H-indazole-3-carboxylic acid [1p-tolyl-meth-(E)-ylidene]-hydrazide (termed as Suprafenacine, SRF). SRF, identified by in silico screening of annotated chemical libraries, was shown to bind microtubules at the colchicine-binding site and inhibit polymerization. This led to G2/M cell cycle arrest and cell death via a mitochondria-mediated apoptotic pathway. Cell death was preceded by loss of mitochondrial membrane potential, JNK - mediated phosphorylation of Bcl-2 and Bad, and activation of caspase-3. Intriguingly, SRF was found to selectively inhibit cancer cell proliferation and was effective against drug-resistant cancer cells by virtue of its ability to bypass the multidrug resistance transporter P-glycoprotein. Taken together, our results suggest that SRF has potential as a chemotherapeutic agent for cancer treatment and provides an alternate scaffold for the development of improved anti-cancer agents. PMID:25354194

Biomass and nitrogen accumulation of hairy vetch-cereal rye cover crop mixtures as influenced by species proportions

USDA-ARS?s Scientific Manuscript database

The performance and suitability of a legume-grass cover crop mixture for specific functions may be influenced by the proportions of each species in the mixture. The objectives of this study were to: 1) evaluate aboveground biomass and species biomass proportions at different hairy vetch (Vicia villo...

Thermodynamics of hairy black holes in Lovelock gravity

NASA Astrophysics Data System (ADS)

Hennigar, Robie A.; Tjoa, Erickson; Mann, Robert B.

2017-02-01

We perform a thorough study of the thermodynamic properties of a class of Lovelock black holes with conformal scalar hair arising from coupling of a real scalar field to the dimensionally extended Euler densities. We study the linearized equations of motion of the theory and describe constraints under which the theory is free from ghosts/tachyons. We then consider, within the context of black hole chemistry, the thermodynamics of the hairy black holes in the Gauss-Bonnet and cubic Lovelock theories. We clarify the connection between isolated critical points and thermodynamic singularities, finding a one parameter family of these critical points which occur for well-defined thermodynamic parameters. We also report on a number of novel results, including `virtual triple points' and the first example of a `λ-line' — a line of second order phase transitions — in black hole thermodynamics.

Effect of anti-glycolytic agents on tumour cells in vitro

NASA Astrophysics Data System (ADS)

Korshunov, D. A.; Kondakova, I. V.

2016-08-01

A metabolic change is one of the tumour hallmarks, which has recently attracted a great amount of attention. One of the main metabolic characteristics of tumour cells is a high level of glycolysis even in the presence of oxygen, known as aerobic glycolysis or the Warburg effect. The energy production is much less in a glycolysis pathway than that in a tricarboxylic acid cycle. The Warburg effect constitutes a fundamental adaptation of tumour cells to a relatively hostile environment, and supports the evolution of aggressive and metastatic phenotypes. As a result, tumour glycolysis may become an attractive target for cancer therapy. Here, we research the effect of potential anticancer agents on tumour cells in vitro. In our study, we found a high sensitivity of tumour cells to anti-glycolityc drugs. In addition, tumour cells are more resistant to the agents studied in comparison with normal cells. We also observed an atypical cooperative interaction of tumour cells in the median lethal dose of drugs. They formed the specific morphological structure of the surviving cells. This behavior is not natural for the culture of tumour cells. Perhaps this is one of the mechanisms of cells' adaptation to the aggressive environment.

78 FR 59099 - Agency Information Collection (Disability Benefits Questionnaires) Under OMB Review

Federal Register 2010, 2011, 2012, 2013, 2014

2013-09-25

.... Hairy Cell and Other B-Cell Leukemias Disability Benefits Questionnaire, VA Form 21-0960b-1. c... exposure: Hairy Cell and Other Chronic B-cell Leukemias, Parkinson's and Ischemic Heart diseases. Veterans...-1--13,750. b. Hairy Cell and Other B-Cell Leukemias Disability Benefits Questionnaire, VA Form 21...

Oxidative phosphorylation-dependent regulation of cancer cell apoptosis in response to anticancer agents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yadav, N.; Kumar, S.; Marlowe, T.

Cancer cells tend to develop resistance to various types of anticancer agents, whether they adopt similar or distinct mechanisms to evade cell death in response to a broad spectrum of cancer therapeutics is not fully defined. Current study concludes that DNA-damaging agents (etoposide and doxorubicin), ER stressor (thapsigargin), and histone deacetylase inhibitor (apicidin) target oxidative phosphorylation (OXPHOS) for apoptosis induction, whereas other anticancer agents including staurosporine, taxol, and sorafenib induce apoptosis in an OXPHOS-independent manner. DNA-damaging agents promoted mitochondrial biogenesis accompanied by increased accumulation of cellular and mitochondrial ROS, mitochondrial protein-folding machinery, and mitochondrial unfolded protein response. Induction of mitochondrialmore » biogenesis occurred in a caspase activation-independent mechanism but was reduced by autophagy inhibition and p53-deficiency. Abrogation of complex-I blocked DNA-damage-induced caspase activation and apoptosis, whereas inhibition of complex-II or a combined deficiency of OXPHOS complexes I, III, IV, and V due to impaired mitochondrial protein synthesis did not modulate caspase activity. Mechanistic analysis revealed that inhibition of caspase activation in response to anticancer agents associates with decreased release of mitochondrial cytochrome c in complex-I-deficient cells compared with wild type (WT) cells. Gross OXPHOS deficiencies promoted increased release of apoptosis-inducing factor from mitochondria compared with WT or complex-I-deficient cells, suggesting that cells harboring defective OXPHOS trigger caspase-dependent as well as caspase-independent apoptosis in response to anticancer agents. Interestingly, DNA-damaging agent doxorubicin showed strong binding to mitochondria, which was disrupted by complex-I-deficiency but not by complex-II-deficiency. Thapsigargin-induced caspase activation was reduced upon abrogation of complex-I or gross OXPHOS

Oxidative phosphorylation-dependent regulation of cancer cell apoptosis in response to anticancer agents.

PubMed

Yadav, N; Kumar, S; Marlowe, T; Chaudhary, A K; Kumar, R; Wang, J; O'Malley, J; Boland, P M; Jayanthi, S; Kumar, T K S; Yadava, N; Chandra, D

2015-11-05

Cancer cells tend to develop resistance to various types of anticancer agents, whether they adopt similar or distinct mechanisms to evade cell death in response to a broad spectrum of cancer therapeutics is not fully defined. Current study concludes that DNA-damaging agents (etoposide and doxorubicin), ER stressor (thapsigargin), and histone deacetylase inhibitor (apicidin) target oxidative phosphorylation (OXPHOS) for apoptosis induction, whereas other anticancer agents including staurosporine, taxol, and sorafenib induce apoptosis in an OXPHOS-independent manner. DNA-damaging agents promoted mitochondrial biogenesis accompanied by increased accumulation of cellular and mitochondrial ROS, mitochondrial protein-folding machinery, and mitochondrial unfolded protein response. Induction of mitochondrial biogenesis occurred in a caspase activation-independent mechanism but was reduced by autophagy inhibition and p53-deficiency. Abrogation of complex-I blocked DNA-damage-induced caspase activation and apoptosis, whereas inhibition of complex-II or a combined deficiency of OXPHOS complexes I, III, IV, and V due to impaired mitochondrial protein synthesis did not modulate caspase activity. Mechanistic analysis revealed that inhibition of caspase activation in response to anticancer agents associates with decreased release of mitochondrial cytochrome c in complex-I-deficient cells compared with wild type (WT) cells. Gross OXPHOS deficiencies promoted increased release of apoptosis-inducing factor from mitochondria compared with WT or complex-I-deficient cells, suggesting that cells harboring defective OXPHOS trigger caspase-dependent as well as caspase-independent apoptosis in response to anticancer agents. Interestingly, DNA-damaging agent doxorubicin showed strong binding to mitochondria, which was disrupted by complex-I-deficiency but not by complex-II-deficiency. Thapsigargin-induced caspase activation was reduced upon abrogation of complex-I or gross OXPHOS deficiency

Oxidative phosphorylation-dependent regulation of cancer cell apoptosis in response to anticancer agents

DOE PAGES

Yadav, N.; Kumar, S.; Marlowe, T.; ...

2015-11-05

Cancer cells tend to develop resistance to various types of anticancer agents, whether they adopt similar or distinct mechanisms to evade cell death in response to a broad spectrum of cancer therapeutics is not fully defined. Current study concludes that DNA-damaging agents (etoposide and doxorubicin), ER stressor (thapsigargin), and histone deacetylase inhibitor (apicidin) target oxidative phosphorylation (OXPHOS) for apoptosis induction, whereas other anticancer agents including staurosporine, taxol, and sorafenib induce apoptosis in an OXPHOS-independent manner. DNA-damaging agents promoted mitochondrial biogenesis accompanied by increased accumulation of cellular and mitochondrial ROS, mitochondrial protein-folding machinery, and mitochondrial unfolded protein response. Induction of mitochondrialmore » biogenesis occurred in a caspase activation-independent mechanism but was reduced by autophagy inhibition and p53-deficiency. Abrogation of complex-I blocked DNA-damage-induced caspase activation and apoptosis, whereas inhibition of complex-II or a combined deficiency of OXPHOS complexes I, III, IV, and V due to impaired mitochondrial protein synthesis did not modulate caspase activity. Mechanistic analysis revealed that inhibition of caspase activation in response to anticancer agents associates with decreased release of mitochondrial cytochrome c in complex-I-deficient cells compared with wild type (WT) cells. Gross OXPHOS deficiencies promoted increased release of apoptosis-inducing factor from mitochondria compared with WT or complex-I-deficient cells, suggesting that cells harboring defective OXPHOS trigger caspase-dependent as well as caspase-independent apoptosis in response to anticancer agents. Interestingly, DNA-damaging agent doxorubicin showed strong binding to mitochondria, which was disrupted by complex-I-deficiency but not by complex-II-deficiency. Thapsigargin-induced caspase activation was reduced upon abrogation of complex-I or gross OXPHOS

Congenital hairy polyp of the oropharynx presenting as an esophageal mass in a neonate, a case report and literature review.

PubMed

Richter, Amy; Mysore, Krupa; Schady, Deb; Chandy, Binoy

2016-01-01

To review the literature of congenital hairy polyps and describe the clinical presentation, operative management, and histologic findings of a congenital hairy polyp arising from the palatopharyngeus muscle in a neonate with recurrent choking episodes. Chart review of a 2-month-old male referred to a tertiary care pediatric hospital. We present a case of a 2-month-old male who presented to the emergency room with recurrent episodes of choking and vomiting. The patient was previously healthy with no prior medical or neonatal history. The parents noted a small fleshy mass in the patient's oropharynx that he would chew on and swallow after several minutes. However, on physical exam, there was no evidence of oropharyngeal mass. The patient did not have respiratory distress. Imaging revealed a 22×7×11mm oblong, fatty mass in the lower cervical and upper thoracic esophagus with a thin stalk extending proximally to the upper collapsed esophagus. Intraoperative recorded laryngoscopy revealed a pedunculated soft palate mass attached to the right superior palatopharyngeus muscle. Histopathology revealed ectodermal and mesodermal elements in a polypoid structure lined by keratinizing squamous epithelium with adnexal structures and central mature adipose tissue, consistent with congenital hairy polyp resembling an accessory tragus of the ear and branchial anomaly. At 6-week follow up, the patient was doing well and gaining weight appropriately with no further choking episodes. There was no evidence of velopharyngeal dysfunction on follow up exam. The surgical site was completely healed and there was no evidence of recurrence. Congenital hairy polyps of the naso- and oropharynx are rare but may present as airway or esophageal masses, causing respiratory distress or choking episodes in a pediatric patient. The pathologic findings of keratinizing squamous epithelium, adnexal structures, adipose and cartilage tissues resemble congenital accessory tragus and may be considered a

Determination of escin content in androgenic embryos and hairy root culture of Aesculus hippocastanum.

PubMed

Calić-Dragosavac, Dusica; Zdravković-Korać, Snezana; Savikin-Fodulović, Katarina; Radojević, Ljiljana; Vinterhalter, Branka

2010-05-01

Escin, a group of chemically related triterpenic glycosides, is widely used in commercial preparations for the treatment of venous insufficiency. Since the zygotic embryo cotyledons accumulate the highest amount of escin, it is currently extracted from the seeds of horse chestnut, Aesculus hippocastanum L. (Hippocastanaceae), on a large scale. As this material is available during only short period of the year, we studied the possibility of using plant tissue culture to obtain escin. For this purpose, the content of escin in androgenic embryos and hairy root cultures of horse chestnut was studied. Escin content was found to be dependent on the stage of androgenic embryo development and the type of phytoregulator supplemented to the nutritive medium. In the absence of phytoregulators, androgenic embryos at the globular stage of development contained approximately four times less escin than those at the cotyledonary stage. Inclusion of various phytoregulators in the nutritive media stimulated escin production. Among them, 2,4-dichlorophenoxyacetic acid (2,4-D) showed the most pronounced effect, with escin content almost reaching that found in zygotic embryos (6.77% versus 6.96%). Two hairy root clones produced substantial amounts of escin (3.57% and 4.09%), less than zygotic embryos, but higher than cotyledonary embryos on phytoregulator-free medium.

Thermodynamical properties of hairy black holes in n spacetime dimensions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nadalini, Mario; Vanzo, Luciano; Zerbini, Sergio

The issue concerning the existence of exact black hole solutions in the presence of a nonvanishing cosmological constant and scalar fields is reconsidered. With regard to this, in investigating no-hair theorem violations, exact solutions of gravity having as a source an interacting and conformally coupled scalar field are revisited in arbitrary dimensional nonasymptotically flat space-times. New and known hairy black hole solutions are discussed. The thermodynamical properties associated with these solutions are investigated and the invariance of the black hole entropy with respect to different conformal frames is proved. The issue of the positivity of the entropy is discussed andmore » resolved for the case of black holes immersed in de Sitter space.« less

DNA excision repair in cell extracts from human cell lines exhibiting hypersensitivity to DNA-damaging agents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hansson, J.; Keyse, S.M.; Lindahl, T.

Whole cell extracts from human lymphoid cell lines can perform in vitro DNA repair synthesis in plasmids damaged by agents including UV or cis-diamminedichloroplatinum(II) (cis-DDP). Extracts from xeroderma pigmentosum (XP) cells are defective in repair synthesis. We have now studied in vitro DNA repair synthesis using extracts from lymphoblastoid cell lines representing four human hereditary syndromes with increased sensitivity to DNA-damaging agents. Extracts of cell lines from individuals with the sunlight-sensitive disorders dysplastic nevus syndrome or Cockayne's syndrome (complementation groups A and B) showed normal DNA repair synthesis in plasmids with UV photoproducts. This is consistent with in vivo measurementsmore » of the overall DNA repair capacity in such cell lines. A number of extracts were prepared from two cell lines representing the variant form of XP (XP-V). Half of the extracts prepared showed normal levels of in vitro DNA repair synthesis in plasmids containing UV lesions, but the remainder of the extracts from the same cell lines showed deficient repair synthesis, suggesting the possibility of an unusually labile excision repair protein in XP-V. Fanconi's anemia (FA) cells show cellular hypersensitivity to cross-linking agents including cis-DDP. Extracts from cell lines belonging to two different complementation groups of FA showed normal DNA repair synthesis in plasmids containing cis-DDP or UV adducts. Thus, there does not appear to be an overall excision repair defect in FA, but the data do not exclude a defect in the repair of interstrand DNA cross-links.« less

Role of pectolytic enzymes in the programmed separation of cells from the root cap of higher plants. Final report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hawes, M.C.

1995-03-01

The objective of this research was to develop a model system to study border cell separation in transgenic pea roots. In addition, the hypothesis that genes encoding pectolytic enzymes in the root cap play a role in the programmed separation of root border cells from the root tip was tested. The following objectives have been accomplished: (1) the use of transgenic hairy roots to study border cell separation has been optimized for Pisum sativum; (2) a cDNA encoding a root cap pectinmethylesterase (PME) has been cloned; (3) PME and polygalacturonase activities in cell walls of the root cap have beenmore » characterized and shown to be correlated with border cell separation. A fusion gene encoding pectate lyase has also been transformed into pea hairy root cells.« less

Reassessment of the hairy long-nosed armadillo "Dasypus" pilosus (Xenarthra, Dasypodidae) and revalidation of the genus Cryptophractus Fitzinger, 1856.

PubMed

Castro, Mariela C; Ciancio, Martín R; Pacheco, Víctor; Salas-Gismondi, Rodolfo M; Bostelmann, J Enrique; Carlini, Alfredo A

2015-04-14

The hairy long-nosed armadillo, currently referred as Dasypus (Cryptophractus) pilosus, is an enigmatic species endemic to montane cloud forests and subparamo of Peruvian Andes. Its strikingly different external features, which include the carapace concealed by abundant hair, the presence of more movable bands, and a slender skull, have raised questions regarding its taxonomic status as subgenus or as genus. This paper assesses this issue based on a cladistic study and provides a detailed comparative description of the species, including the first account on the distinctive ornamentation of its osteoderms. Based on several unique characters in the carapace, skull, mandible, and teeth, as well as on the external phylogenetic position relative to other Dasypus, we favor the assignment of the hairy long-nosed armadillo to other genus. As result, we revalidate the original generic epithet, so that the valid name of the species is Cryptophractus pilosus Fitzinger, 1856.

Combining engineered cell-sensors with multi-agent systems to realize smart environment

NASA Astrophysics Data System (ADS)

Chen, Mei

2013-03-01

The connection of everything in a sensory and an intelligent way is a pursuit in smart environment. This paper introduces the engineered cell-sensors into the multi-agent systems to realize the smart environment. The seamless interface with the natural environment and strong information-processing ability of cell with the achievements of synthetic biology make the construction of engineered cell-sensors possible. However, the engineered cell-sensors are only simple-functional and unreliable computational entities. Therefore how to combine engineered cell-sensors with digital device is a key problem in order to realize the smart environment. We give the abstract structure and interaction modes of the engineered cell-sensors in order to introduce engineered cell-sensors into multi-agent systems. We believe that the introduction of engineered cell-sensors will push forward the development of the smart environment.

Molecular characteristics and efficacy of 16D10 siRNAs in inhibiting root-knot nematode infection in transgenic grape hairy roots.

PubMed

Yang, Yingzhen; Jittayasothorn, Yingyos; Chronis, Demosthenis; Wang, Xiaohong; Cousins, Peter; Zhong, Gan-Yuan

2013-01-01

Root-knot nematodes (RKNs) infect many annual and perennial crops and are the most devastating soil-born pests in vineyards. To develop a biotech-based solution for controlling RKNs in grapes, we evaluated the efficacy of plant-derived RNA interference (RNAi) silencing of a conserved RKN effector gene, 16D10, for nematode resistance in transgenic grape hairy roots. Two hairpin-based silencing constructs, containing a stem sequence of 42 bp (pART27-42) or 271 bp (pART27-271) of the 16D10 gene, were transformed into grape hairy roots and compared for their small interfering RNA (siRNA) production and efficacy on suppression of nematode infection. Transgenic hairy root lines carrying either of the two RNAi constructs showed less susceptibility to nematode infection compared with control. Small RNA libraries from four pART27-42 and two pART27-271 hairy root lines were sequenced using an Illumina sequencing technology. The pART27-42 lines produced hundred times more 16D10-specific siRNAs than the pART27-271 lines. On average the 16D10 siRNA population had higher GC content than the 16D10 stem sequences in the RNAi constructs, supporting previous observation that plant dicer-like enzymes prefer GC-rich sequences as substrates for siRNA production. The stems of the 16D10 RNAi constructs were not equally processed into siRNAs. Several hot spots for siRNA production were found in similar positions of the hairpin stems in pART27-42 and pART27-271. Interestingly, stem sequences at the loop terminus produced more siRNAs than those at the stem base. Furthermore, the relative abundance of guide and passenger single-stranded RNAs from putative siRNA duplexes was largely correlated with their 5' end thermodynamic strength. This study demonstrated the feasibility of using a plant-derived RNAi approach for generation of novel nematode resistance in grapes and revealed several interesting molecular characteristics of transgene siRNAs important for optimizing plant RNAi constructs.

Doxorubicin Delivery into Tumor Cells by Stable Cavitation without Contrast Agents.

PubMed

Chettab, Kamel; Mestas, Jean-Louis; Lafond, Maxime; Saadna, Djamel Eddine; Lafon, Cyril; Dumontet, Charles

2017-02-06

Doxorubicin, alone or in combination with other anticancer agents, is one of the most widely used chemotherapeutic agents and is administered in a wide range of cancers. However, the use of doxorubicin is limited due to its potential serious adverse reactions. Previous studies have established the ability of high intensity focused ultrasound (HIFU) in combination with various contrast agents to increase intracellular doxorubicin delivery in a targeted and noninvasive manner. In this study, we developed a new sonoporation device generating and monitoring acoustic cavitation bubbles without any addition of contrast agents. The device was used to potentiate the delivery of active doxorubicin into both adherent and suspended cell lines. Combining doxorubicin with ultrasound resulted in a significant enhancement of doxorubicin intracellular delivery and a decrease in cell viability at 48 and 72 h, in comparison to doxorubicin alone. More importantly and unlike previous investigations, our procedure does not require the addition of contrast agents to generate acoustic cavitation and to achieve high levels of doxorubicin delivery. The successful translation of this approach for an in vivo application may allow a significant reduction in the dosage and the adverse effects of doxorubicin therapy in patients.

Cytotoxic Effects of the Radiocontrast Agent Iotrolan and Anesthetic Agents Bupivacaine and Lidocaine in Three-Dimensional Cultures of Human Intervertebral Disc Nucleus Pulposus Cells: Identification of the Apoptotic Pathways

PubMed Central

Iwasaki, Koji; Sudo, Hideki; Yamada, Katsuhisa; Ito, Manabu; Iwasaki, Norimasa

2014-01-01

Background Discography and discoblock are imaging procedures used to diagnose discogenic low back pain. Although needle puncture of the intervertebral disc (IVD) itself induces disc degeneration, the agents used in these procedures may also have harmful effects on IVD cells. The purpose of this study was to analyze whether radiocontrast agents and local anesthetic agents have detrimental effects on human nucleus pulposus (NP) cells. Methods Healthy human NP cells were cultured for 7 days in three-dimensional (3D) cell–alginate bead composites, and were then exposed to clinically relevant doses of a radiocontrast agent (iotrolan) or local anesthetic (lidocaine or bupivacaine). Cell viability and apoptosis were measured by confocal microscopy and flow cytometry. On the basis of caspase expression profiles, the apoptotic pathways activated by the agents were identified by Western blot analysis. Results The radiocontrast agent iotrolan did not affect NP cell viability or induce apoptosis. In contrast, both the anesthetic agents significantly decreased cell viability and increased the apoptotic cell number in a time- and dose-dependent manner. After 120 min, 2% lidocaine and 0.5% bupivacaine decreased percent live cells to 13% and 10%, respectively (p<0.05). The number of apoptotic cells was doubled by increasing lidocaine dosage from 1% to 2% (23% and 42%) and bupivacaine from 0.25% to 0.50% (25% and 48%) (p<0.05). Western blot analysis revealed that both anesthetic agents upregulated cleaved caspase-3 and caspase-8, whereas only bupivacaine upregulated cleaved caspase-9. Conclusions/Significance The present study demonstrates that iotrolan does not affect the viability of healthy human NP cells. In contrast, the two anesthetic agents commonly used in discography or discoblock may cause extensive damage to IVDs by inducing apoptotic cell death. PMID:24642945

Genomic cloning and chromosomal localization of HRY, the human homolog to the Drosophila segmentation gene, hairy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Feder, J.N.; Jan, L.Y.; Jan, Y.N.

The Drosophila hairy gene encodes a basic helix- loop-helix protein that functions in at least two steps during Drosophila development: (1) during embryogenesis, when it partakes in the establishment of segments, and (2) during the larval stage, when it functions negatively in determining the pattern of sensory bristles on the adult fly. In the rat, a structurally homologous gene (RHL) behaves as an immediate-early gene in its response to growth factors and can, like that in Drosophila, suppress neuronal differentiation events. Here, the authors report the genomic cloning of the human hairy gene homolog (HRY). The coding region of themore » gene is contained within four exons. The predicted amino acid sequence reveals only four amino acid differences between the human and rat genes. Analysis of the DNA sequence 5[prime] to the coding region reveals a putatitve untranslated exon. To increase the value of the HRY gene as a genetic marker and to assess its potential involvement in genetic disorders, they sublocalized the locus to chromosome 3q28-q29 by fluorescence in situ hybridization. 34 refs., 4 figs., 1 tab.« less

Hairy vetch biomass across the eastern United States: Effects of latitude, seeding rate and date, and termination timing

USDA-ARS?s Scientific Manuscript database

Hairy vetch (Vicia villosa Roth) is a winter annual legume cover crop that is often grown because it can provide a substantial amount of N to the following cash crop. Nitrogen accumulation is dependent on biomass production, which in turn is affected by climate, seeding rate and date, and timing of ...

Hairy roots of Helianthus annuus: a model system to study phytoremediation of tetracycline and oxytetracycline.

PubMed

Gujarathi, Ninad P; Haney, Bryan J; Park, Heidi J; Wickramasinghe, S Ranil; Linden, James C

2005-01-01

The release of antibiotics to the environment has to be controlled because of serious threats to human health. Hairy root cultures of Helianthus annuus (sunflower), along with their inherent rhizospheric activity, provide a fast growing, microbe-free environment for understanding plant-pollutant interactions. The root system catalyzes rapid disappearance of tetracycline (TC) and oxytetracycline (OTC) from aqueous media, which suggests roots have potential for phytoremediation of the two antibiotics in vivo. In addition, in vitro modifications of the two antibiotics by filtered, cell- and microbe-free root exudates suggest involvement of root-secreted compounds. The modification is confirmed from changes observed in UV spectra of exudate-treated OTC. Modification appears to be more dominant at the BCD chromophore of the antibiotic molecule. Kinetic analyses dismiss direct enzyme catalysis; the modification rates decrease with increasing OTC concentrations. The rates increase with increasing age of cultures from which root exudates are prepared. The decrease in modification rates upon addition of the antioxidant ascorbic acid (AA) suggests involvement of reactive oxygen species (ROS) in the antibiotic modification process.

The Essential Oils of Rhaponticum carthamoides Hairy Roots and Roots of Soil-Grown Plants: Chemical Composition and Antimicrobial, Anti-Inflammatory, and Antioxidant Activities.

PubMed

Skała, Ewa; Rijo, Patrícia; Garcia, Catarina; Sitarek, Przemysław; Kalemba, Danuta; Toma, Monika; Szemraj, Janusz; Pytel, Dariusz; Wysokińska, Halina; Śliwiński, Tomasz

2016-01-01

The essential oils were isolated by hydrodistillation from the hairy roots (HR) and roots of soil-grown plants (SGR) of Rhaponticum carthamoides and were analyzed by GC-MS method. In the both essential oils 62 compounds were identified. The root essential oils showed the differences in the qualitative and quantitative composition. The sesquiterpene hydrocarbons (55-62%) dominated in both essential oils. The major compounds of HR essential oil were cyperene, 13-norcypera-1(5),11(12)-diene, and cadalene while aplotaxene, nardosina-1(10),11-diene, and dauca-4(11),8-diene dominated in SGR essential oil. Both essential oils showed antibacterial activity especially against Enterococcus faecalis (ATCC 29212) and Pseudomonas aeruginosa (ATCC 27853) (MIC value = 125  µ g/mL). HR and SGR essential oils also decreased the expression of IL-1 β , IL-6, and TNF- α and the ROS level in LPS-treatment astrocytes. This is the first report to describe the chemical composition of R. carthamoides essential oil from hairy roots, its protective effect against LPS-induced inflammation and ROS production in astrocytes, and its antimicrobial potential. The results show that R. carthamoides hairy roots may be a valuable source of the essential oil and may be an alternative to the roots of soil-grown plants.

The Essential Oils of Rhaponticum carthamoides Hairy Roots and Roots of Soil-Grown Plants: Chemical Composition and Antimicrobial, Anti-Inflammatory, and Antioxidant Activities

PubMed Central

Rijo, Patrícia; Garcia, Catarina; Kalemba, Danuta; Toma, Monika; Szemraj, Janusz; Pytel, Dariusz; Śliwiński, Tomasz

2016-01-01

The essential oils were isolated by hydrodistillation from the hairy roots (HR) and roots of soil-grown plants (SGR) of Rhaponticum carthamoides and were analyzed by GC-MS method. In the both essential oils 62 compounds were identified. The root essential oils showed the differences in the qualitative and quantitative composition. The sesquiterpene hydrocarbons (55–62%) dominated in both essential oils. The major compounds of HR essential oil were cyperene, 13-norcypera-1(5),11(12)-diene, and cadalene while aplotaxene, nardosina-1(10),11-diene, and dauca-4(11),8-diene dominated in SGR essential oil. Both essential oils showed antibacterial activity especially against Enterococcus faecalis (ATCC 29212) and Pseudomonas aeruginosa (ATCC 27853) (MIC value = 125 µg/mL). HR and SGR essential oils also decreased the expression of IL-1β, IL-6, and TNF-α and the ROS level in LPS-treatment astrocytes. This is the first report to describe the chemical composition of R. carthamoides essential oil from hairy roots, its protective effect against LPS-induced inflammation and ROS production in astrocytes, and its antimicrobial potential. The results show that R. carthamoides hairy roots may be a valuable source of the essential oil and may be an alternative to the roots of soil-grown plants. PMID:28074117

Sensitization of gastric cancer cells to alkylating agents by glaucocalyxin B via cell cycle arrest and enhanced cell death

PubMed Central

Ur Rahman, Muhammad Saif; Zhang, Ling; Wu, Lingyan; Xie, Yuqiong; Li, Chunchun; Cao, Jiang

2017-01-01

Severe side effects are major problems with chemotherapy of gastric cancer (GC). These side effects can be reduced by using sensitizing agents in combination with therapeutic drugs. In this study, the low/nontoxic dosage of glaucocalyxin B (GLB) was used with other DNA linker agents mitomycin C (MMC), cisplatin (DDP), or cyclophosphamide (CTX) to treat GC cells. Combined effectiveness of GLB with drugs was determined by proliferation assay. The molecular mechanisms associated with cell proliferation, migration, invasion, cell cycle, DNA repair/replication, apoptosis, and autophagy were investigated by immunoblotting for key proteins involved. Cell cycle and apoptosis analysis were performed by flow cytometry. Reactive oxygen species level was also examined for identification of its role in apoptosis. Proliferation assay revealed that the addition of 5 µM GLB significantly sensitizes gastric cancer SGC-7901 cells to MMC, DDP, and CTX by decreasing half-maximal inhibitory concentration (IC50) by up to 75.40%±5%, 45.10%±5%, and 52.10%±5%, respectively. GLB + drugs decreased the expression level of proteins involved in proliferation and migration, suggesting the anticancer potential of GLB + drugs. GLB + MMC, GLB + CTX, and GLB + DDP arrest the cells in G0/G1 and G1/S phase, respectively, which may be the consequence of significant decrease in the level of enzymes responsible for DNA replication and telomerase shortening. Combined use of GLB with these drugs also induces DNA damage and apoptosis by activating caspase/PARP pathways and increased production of reactive oxygen species and increased autophagy in GC cells. GLB dosage sensitizes GC cells to the alkylating agents via arresting the cell cycle and enhancing cell death. This is of significant therapeutic importance in the reduction of side effects associated with these drugs. PMID:28860714

Sensitization of gastric cancer cells to alkylating agents by glaucocalyxin B via cell cycle arrest and enhanced cell death.

PubMed

Ur Rahman, Muhammad Saif; Zhang, Ling; Wu, Lingyan; Xie, Yuqiong; Li, Chunchun; Cao, Jiang

2017-01-01

Severe side effects are major problems with chemotherapy of gastric cancer (GC). These side effects can be reduced by using sensitizing agents in combination with therapeutic drugs. In this study, the low/nontoxic dosage of glaucocalyxin B (GLB) was used with other DNA linker agents mitomycin C (MMC), cisplatin (DDP), or cyclophosphamide (CTX) to treat GC cells. Combined effectiveness of GLB with drugs was determined by proliferation assay. The molecular mechanisms associated with cell proliferation, migration, invasion, cell cycle, DNA repair/replication, apoptosis, and autophagy were investigated by immunoblotting for key proteins involved. Cell cycle and apoptosis analysis were performed by flow cytometry. Reactive oxygen species level was also examined for identification of its role in apoptosis. Proliferation assay revealed that the addition of 5 µM GLB significantly sensitizes gastric cancer SGC-7901 cells to MMC, DDP, and CTX by decreasing half-maximal inhibitory concentration (IC 50 ) by up to 75.40%±5%, 45.10%±5%, and 52.10%±5%, respectively. GLB + drugs decreased the expression level of proteins involved in proliferation and migration, suggesting the anticancer potential of GLB + drugs. GLB + MMC, GLB + CTX, and GLB + DDP arrest the cells in G 0 /G 1 and G 1 /S phase, respectively, which may be the consequence of significant decrease in the level of enzymes responsible for DNA replication and telomerase shortening. Combined use of GLB with these drugs also induces DNA damage and apoptosis by activating caspase/PARP pathways and increased production of reactive oxygen species and increased autophagy in GC cells. GLB dosage sensitizes GC cells to the alkylating agents via arresting the cell cycle and enhancing cell death. This is of significant therapeutic importance in the reduction of side effects associated with these drugs.

Preclinical Investigations of PM01183 (Lurbinectedin) as a Single Agent or in Combination with Other Anticancer Agents for Clear Cell Carcinoma of the Ovary

PubMed Central

Takahashi, Ryoko; Mabuchi, Seiji; Kawano, Mahiru; Sasano, Tomoyuki; Matsumoto, Yuri; Kuroda, Hiromasa; Kozasa, Katsumi; Hashimoto, Kae; Sawada, Kenjiro; Kimura, Tadashi

2016-01-01

Objective The objective of this study was to evaluate the antitumor effects of lurbinectedin as a single agent or in combination with existing anticancer agents for clear cell carcinoma (CCC) of the ovary, which is regarded as an aggressive, chemoresistant, histological subtype. Methods Using human ovarian CCC cell lines, the antitumor effects of lurbinectedin, SN-38, doxorubicin, cisplatin, and paclitaxel as single agents were assessed using the MTS assay. Then, the antitumor effects of combination therapies involving lurbinectedin and 1 of the other 4 agents were evaluated using isobologram analysis to examine whether these combinations displayed synergistic effects. The antitumor activity of each treatment was also examined using cisplatin-resistant and paclitaxel-resistant CCC sublines. Finally, we determined the effects of mTORC1 inhibition on the antitumor activity of lurbinectedin-based chemotherapy. Results Lurbinectedin exhibited significant antitumor activity toward chemosensitive and chemoresistant CCC cells in vitro. An examination of mouse CCC cell xenografts revealed that lurbinectedin significantly inhibits tumor growth. Among the tested combinations, lurbinectedin plus SN-38 resulted in a significant synergistic effect. This combination also had strong synergistic effects on both the cisplatin-resistant and paclitaxel-resistant CCC cell lines. Everolimus significantly enhanced the antitumor activity of lurbinectedin-based chemotherapies. Conclusions Lurbinectedin, a new agent that targets active transcription, exhibits antitumor activity in CCC when used as a single agent and has synergistic antitumor effects when combined with irinotecan. Our results indicate that lurbinectedin is a promising agent for treating ovarian CCC, both as a first-line treatment and as a salvage treatment for recurrent lesions that develop after platinum-based or paclitaxel treatment. PMID:26986199

High-speed homogenization coupled with microwave-assisted extraction followed by liquid chromatography-tandem mass spectrometry for the direct determination of alkaloids and flavonoids in fresh Isatis tinctoria L. hairy root cultures.

PubMed

Jiao, Jiao; Gai, Qing-Yan; Zhang, Lin; Wang, Wei; Luo, Meng; Zu, Yuan-Gang; Fu, Yu-Jie

2015-06-01

A new, simple and efficient analysis method for fresh plant in vitro cultures-namely, high-speed homogenization coupled with microwave-assisted extraction (HSH-MAE) followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS)-was developed for simultaneous determination of six alkaloids and eight flavonoids in Isatis tinctoria hairy root cultures (ITHRCs). Compared with traditional methods, the proposed HSH-MAE offers the advantages of easy manipulation, higher efficiency, energy saving, and reduced waste. Cytohistological studies were conducted to clarify the mechanism of HSH-MAE at cellular/tissue levels. Moreover, the established LC-MS/MS method showed excellent linearity, precision, repeatability, and reproducibility. The HSH-MAE-LC-MS/MS method was also successfully applied for screening high-productivity ITHRCs. Overall, this study opened up a new avenue for the direct determination of secondary metabolic profiles from fresh plant in vitro cultures, which is valuable for improving quality control of plant cell/organ cultures and sheds light on the metabolomic analysis of biological samples. Graphical Abstract HSH-MAE-LC-MS/MS opened up a new avenue for the direct determination of alkaloids and flavonoids in fresh Isatis tinctoria hairy root cultures.

Multicenter retrospective analysis regarding the clinical manifestations and treatment results in patients with hairy cell leukemia: twenty-four year Turkish experience in cladribine therapy.

PubMed

Hacioglu, Sibel; Bilen, Yusuf; Eser, Ali; Sivgin, Serdar; Gurkan, Emel; Yildirim, Rahsan; Aydogdu, Ismet; Dogu, Mehmet Hilmi; Yilmaz, Mehmet; Kayikci, Omur; Tombak, Anil; Kuku, Irfan; Celebi, Harika; Akay, Meltem Olga; Esen, Ramazan; Korkmaz, Serdal; Keskin, Ali

2015-12-01

In this multicenter retrospective analysis, we aimed to present clinical, laboratory and treatment results of 94 patients with Hairy cell leukemia diagnosed in 13 centers between 1990 and 2014. Sixty-six of the patients were males and 28 were females, with a median age of 55. Splenomegaly was present in 93.5% of cases at diagnosis. The laboratory findings that came into prominence were pancytopenia with grade 3 bone marrow fibrosis. Most of the patients with an indication for treatment were treated with cladribine as first-line treatment. Total and complete response of cladribine was 97.3% and 80.7%. The relapse rate after cladribine was 16.6%, and treatment related mortality was 2.5%. Most preferred therapy (95%) was again cladribine at second-line, and third line with CR rate of 68.4% and 66.6%, respectively. The 28-month median OS was 91.7% in all patients and 25-month median OS 96% for patients who were given cladribine as first-line therapy. In conclusion, the first multicenter retrospective Turkish study where patients with HCL were followed up for a long period has revealed demographic characteristics of patients with HCL, and confirmed that cladribine treatment might be safe and effective in a relatively large series of the Turkish study population. Copyright © 2014 John Wiley & Sons, Ltd.

Influences of Agrobacterium rhizogenes strains, plant genotypes, and tissue types on the induction of transgenic hairy roots in Vitis species

USDA-ARS?s Scientific Manuscript database

Agrobacterium rhizogenes-mediated induction of transgenic hairy roots was previously demonstrated in Vitis vinifera L. and a few other Vitis species. In this study, 13 Vitis species, including V. aestivalis, V. afghanistan, V. champinii, V. doaniana, V. flexuosa, V. labrusca, V. nesbittiana, V. pal...

Holographic Van der Waals phase transition of the higher-dimensional electrically charged hairy black hole

NASA Astrophysics Data System (ADS)

Li, Hui-Ling; Feng, Zhong-Wen; Zu, Xiao-Tao

2018-01-01

With motivation by holography, employing black hole entropy, two-point connection function and entanglement entropy, we show that, for the higher-dimensional Anti-de Sitter charged hairy black hole in the fixed charged ensemble, a Van der Waals-like phase transition can be observed. Furthermore, based on the Maxwell equal-area construction, we check numerically the equal-area law for a first order phase transition in order to further characterize the Van der Waals-like phase transition.

Multiple repair pathways mediate tolerance to chemotherapeutic cross-linking agents in vertebrate cells.

PubMed

Nojima, Kuniharu; Hochegger, Helfrid; Saberi, Alihossein; Fukushima, Toru; Kikuchi, Koji; Yoshimura, Michio; Orelli, Brian J; Bishop, Douglas K; Hirano, Seiki; Ohzeki, Mioko; Ishiai, Masamichi; Yamamoto, Kazuhiko; Takata, Minoru; Arakawa, Hiroshi; Buerstedde, Jean-Marie; Yamazoe, Mitsuyoshi; Kawamoto, Takuo; Araki, Kasumi; Takahashi, Jun A; Hashimoto, Nobuo; Takeda, Shunichi; Sonoda, Eiichiro

2005-12-15

Cross-linking agents that induce DNA interstrand cross-links (ICL) are widely used in anticancer chemotherapy. Yeast genetic studies show that nucleotide excision repair (NER), Rad6/Rad18-dependent postreplication repair, homologous recombination, and cell cycle checkpoint pathway are involved in ICL repair. To study the contribution of DNA damage response pathways in tolerance to cross-linking agents in vertebrates, we made a panel of gene-disrupted clones from chicken DT40 cells, each defective in a particular DNA repair or checkpoint pathway, and measured the sensitivities to cross-linking agents, including cis-diamminedichloroplatinum (II) (cisplatin), mitomycin C, and melphalan. We found that cells harboring defects in translesion DNA synthesis (TLS), Fanconi anemia complementation groups (FANC), or homologous recombination displayed marked hypersensitivity to all the cross-linking agents, whereas NER seemed to play only a minor role. This effect of replication-dependent repair pathways is distinctively different from the situation in yeast, where NER seems to play a major role in dealing with ICL. Cells deficient in Rev3, the catalytic subunit of TLS polymerase Polzeta, showed the highest sensitivity to cisplatin followed by fanc-c. Furthermore, epistasis analysis revealed that these two mutants work in the same pathway. Our genetic comprehensive study reveals a critical role for DNA repair pathways that release DNA replication block at ICLs in cellular tolerance to cross-linking agents and could be directly exploited in designing an effective chemotherapy.

Cloning and expression of UDP-glucose: flavonoid 7-O-glucosyltransferase from hairy root cultures of Scutellaria baicalensis.

PubMed

Hirotani, M; Kuroda, R; Suzuki, H; Yoshikawa, T

2000-05-01

A cDNA encoding UDP-glucose: baicalein 7-O-glucosyltransferase (UBGT) was isolated from a cDNA library from hairy root cultures of Scutellaria baicalensis Georgi probed with a partial-length cDNA clone of a UDP-glucose: flavonoid 3-O-glucosyltransferase (UFGT) from grape (Vitis vinifera L.). The heterologous probe contained a glucosyltransferase consensus amino acid sequence which was also present in the Scutellaria cDNA clones. The complete nucleotide sequence of the 1688-bp cDNA insert was determined and the deduced amino acid sequences are presented. The nucleotide sequence analysis of UBGT revealed an open reading frame encoding a polypeptide of 476 amino acids with a calculated molecular mass of 53,094 Da. The reaction product for baicalein and UDP-glucose catalyzed by recombinant UBGT in Escherichia coli was identified as authentic baicalein 7-O-glucoside using high-performance liquid chromatography and proton nuclear magnetic resonance spectroscopy. The enzyme activities of recombinant UBGT expressed in E. coli were also detected towards flavonoids such as baicalein, wogonin, apigenin, scutellarein, 7,4'-dihydroxyflavone and kaempferol, and phenolic compounds. The accumulation of UBGT mRNA in hairy roots was in response to wounding or salicylic acid treatments.

Cell-type-specific, Aptamer-functionalized Agents for Targeted Disease Therapy

PubMed Central

Zhou, Jiehua; Rossi, John J.

2014-01-01

One hundred years ago, Dr. Paul Ehrlich popularized the “magic bullet” concept for cancer therapy in which an ideal therapeutic agent would only kill the specific tumor cells it targeted. Since then, “targeted therapy” that specifically targets the molecular defects responsible for a patient's condition has become a long-standing goal for treating human disease. However, safe and efficient drug delivery during the treatment of cancer and infectious disease remains a major challenge for clinical translation and the development of new therapies. The advent of SELEX technology has inspired many groundbreaking studies that successfully adapted cell-specific aptamers for targeted delivery of active drug substances in both in vitro and in vivo models. By covalently linking or physically functionalizing the cell-specific aptamers with therapeutic agents, such as siRNA, microRNA, chemotherapeutics or toxins, or delivery vehicles, such as organic or inorganic nanocarriers, the targeted cells and tissues can be specifically recognized and the therapeutic compounds internalized, thereby improving the local concentration of the drug and its therapeutic efficacy. Currently, many cell-type-specific aptamers have been developed that can target distinct diseases or tissues in a cell-type-specific manner. In this review, we discuss recent advances in the use of cell-specific aptamers for targeted disease therapy, as well as conjugation strategies and challenges. PMID:24936916

3′-Phosphoadenosine 5′-phosphosulfate synthase 1 (PAPSS1) knockdown sensitizes non-small cell lung cancer cells to DNA damaging agents

PubMed Central

Leung, Ada W. Y.; Dragowska, Wieslawa H.; Ricaurte, Daniel; Kwok, Brian; Mathew, Veena; Roosendaal, Jeroen; Ahluwalia, Amith; Warburton, Corinna; Laskin, Janessa J.; Stirling, Peter C.; Qadir, Mohammed A.; Bally, Marcel B.

2015-01-01

Standard treatment for advanced non-small cell lung cancer (NSCLC) with no known driver mutation is platinum-based chemotherapy, which has a response rate of only 30–33%. Through an siRNA screen, 3′-phosphoadenosine 5′-phosphosulfate (PAPS) synthase 1 (PAPSS1), an enzyme that synthesizes the biologically active form of sulfate PAPS, was identified as a novel platinum-sensitizing target in NSCLC cells. PAPSS1 knockdown in combination with low-dose (IC10) cisplatin reduces clonogenicity of NSCLC cells by 98.7% (p < 0.001), increases DNA damage, and induces G1/S phase cell cycle arrest and apoptosis. PAPSS1 silencing also sensitized NSCLC cells to other DNA crosslinking agents, radiation, and topoisomerase I inhibitors, but not topoisomerase II inhibitors. Chemo-sensitization was not observed in normal epithelial cells. Knocking out the PAPSS1 homolog did not sensitize yeast to cisplatin, suggesting that sulfate bioavailability for amino acid synthesis is not the cause of sensitization to DNA damaging agents. Rather, sensitization may be due to sulfation reactions involved in blocking the action of DNA damaging agents, facilitating DNA repair, promoting cancer cell survival under therapeutic stress or reducing the bioavailability of DNA damaging agents. Our study demonstrates for the first time that PAPSS1 could be targeted to improve the activity of multiple anticancer agents used to treat NSCLC. PMID:26220590

Antiretroviral Agents Effectively Block HIV Replication after Cell-to-Cell Transfer

PubMed Central

Permanyer, Marc; Ballana, Ester; Ruiz, Alba; Badia, Roger; Riveira-Munoz, Eva; Gonzalo, Encarna; Clotet, Bonaventura

2012-01-01

Cell-to-cell transmission of HIV has been proposed as a mechanism contributing to virus escape to the action of antiretrovirals and a mode of HIV persistence during antiretroviral therapy. Here, cocultures of infected HIV-1 cells with primary CD4+ T cells or lymphoid cells were used to evaluate virus transmission and the effect of known antiretrovirals. Transfer of HIV antigen from infected to uninfected cells was resistant to the reverse transcriptase inhibitors (RTIs) zidovudine (AZT) and tenofovir, but was blocked by the attachment inhibitor IgGb12. However, quantitative measurement of viral DNA production demonstrated that all anti-HIV agents blocked virus replication with similar potency to cell-free virus infections. Cell-free and cell-associated infections were equally sensitive to inhibition of viral replication when HIV-1 long terminal repeat (LTR)-driven green fluorescent protein (GFP) expression in target cells was measured. However, detection of GFP by flow cytometry may incorrectly estimate the efficacy of antiretrovirals in cell-associated virus transmission, due to replication-independent Tat-mediated LTR transactivation as a consequence of cell-to-cell events that did not occur in short-term (48-h) cell-free virus infections. In conclusion, common markers of virus replication may not accurately correlate and measure infectivity or drug efficacy in cell-to-cell virus transmission. When accurately quantified, active drugs blocked proviral DNA and virus replication in cell-to-cell transmission, recapitulating the efficacy of antiretrovirals in cell-free virus infections and in vivo. PMID:22696642

Molecular Characteristics and Efficacy of 16D10 siRNAs in Inhibiting Root-Knot Nematode Infection in Transgenic Grape Hairy Roots

PubMed Central

Chronis, Demosthenis; Wang, Xiaohong; Cousins, Peter; Zhong, Gan-Yuan

2013-01-01

Root-knot nematodes (RKNs) infect many annual and perennial crops and are the most devastating soil-born pests in vineyards. To develop a biotech-based solution for controlling RKNs in grapes, we evaluated the efficacy of plant-derived RNA interference (RNAi) silencing of a conserved RKN effector gene, 16D10, for nematode resistance in transgenic grape hairy roots. Two hairpin-based silencing constructs, containing a stem sequence of 42 bp (pART27-42) or 271 bp (pART27-271) of the 16D10 gene, were transformed into grape hairy roots and compared for their small interfering RNA (siRNA) production and efficacy on suppression of nematode infection. Transgenic hairy root lines carrying either of the two RNAi constructs showed less susceptibility to nematode infection compared with control. Small RNA libraries from four pART27-42 and two pART27-271 hairy root lines were sequenced using an Illumina sequencing technology. The pART27-42 lines produced hundred times more 16D10-specific siRNAs than the pART27-271 lines. On average the 16D10 siRNA population had higher GC content than the 16D10 stem sequences in the RNAi constructs, supporting previous observation that plant dicer-like enzymes prefer GC-rich sequences as substrates for siRNA production. The stems of the 16D10 RNAi constructs were not equally processed into siRNAs. Several hot spots for siRNA production were found in similar positions of the hairpin stems in pART27-42 and pART27-271. Interestingly, stem sequences at the loop terminus produced more siRNAs than those at the stem base. Furthermore, the relative abundance of guide and passenger single-stranded RNAs from putative siRNA duplexes was largely correlated with their 5′ end thermodynamic strength. This study demonstrated the feasibility of using a plant-derived RNAi approach for generation of novel nematode resistance in grapes and revealed several interesting molecular characteristics of transgene siRNAs important for optimizing plant RNAi constructs

Hairy black hole solutions in U(1) gauge-invariant scalar-vector-tensor theories

NASA Astrophysics Data System (ADS)

Heisenberg, Lavinia; Tsujikawa, Shinji

2018-05-01

In U (1) gauge-invariant scalar-vector-tensor theories with second-order equations of motion, we study the properties of black holes (BH) on a static and spherically symmetric background. In shift-symmetric theories invariant under the shift of scalar ϕ → ϕ + c, we show the existence of new hairy BH solutions where a cubic-order scalar-vector interaction gives rise to a scalar hair manifesting itself around the event horizon. In the presence of a quartic-order interaction besides the cubic coupling, there are also regular BH solutions endowed with scalar and vector hairs.

Cladribine in a weekly versus daily schedule for untreated active hairy cell leukemia: final report from the Polish Adult Leukemia Group (PALG) of a prospective, randomized, multicenter trial.

PubMed

Robak, Tadeusz; Jamroziak, Krzysztof; Gora-Tybor, Joanna; Blonski, Jerzy Z; Kasznicki, Marek; Dwilewicz-Trojaczek, Jadwiga; Wiater, Elzbieta; Zdunczyk, Andrzej; Dybowicz, Jacek; Dmoszynska, Anna; Wojtaszko, Maria; Zdziarska, Barbara; Calbecka, Malgorzata; Kostyra, Aleksandra; Hellmann, Andrzej; Lewandowski, Krzysztof; Stella-Holowiecka, Beata; Sulek, Kazimierz; Gawronski, Krzysztof; Skotnicki, Aleksander B; Nowak, Wieslaw; Zawilska, Krystyna; Molendowicz-Portala, Lucyna; Kloczko, Janusz; Sokolowski, Jaroslaw; Warzocha, Krzysztof; Seferynska, Ilona; Ceglarek, Bernardeta; Konopka, Lech

2007-05-01

Cladribine (2-chlorodeoxyadenosine, 2-CdA) treatment-associated infections may shorten potentially long-term survival in hairy cell leukemia (HCL). In search of the optimal mode of 2-CdA administration, 132 patients with untreated HCL were randomized to receive either standard 5-day 2-CdA protocol or a novel schedule of 6 weekly 2-CdA infusions suggested to be less toxic. Analysis of treatment response confirmed similar complete remission rates, overall response rates, progression-free survival, and overall survival in both 2-CdA protocols. However, we did not observe lower toxicity in the weekly schedule. Of special interest, no significant differences were found in the rate of grade 3/4 infections (18% for daily and 26% for weekly protocol, difference -8.2%; 95% confidence interval [CI] -23.2% to 6.9%; P = .28) and the rate of septic deaths (3% for daily and 2% for weekly protocol, difference 1.4%; 95% CI -4.3% to 7.0%; P = .64). In conclusion, HCL treatment with weekly 2-CdA infusions is equally effective but no safer than the standard 5-day 2-CdA protocol.

Sensory Cells of the Fish Ear: A Hairy Enigma

NASA Technical Reports Server (NTRS)

Popper, A. N.; Saidel, W. M.

1995-01-01

Analysis of the structure of the ears in teleost fishes has led to the tentative suggestion that otolithic endorgans may function differently, in different species. Recently, evidence has demonstrated different 'types' of sensory hair cells can be found in the ears of teleost fishes, and individual hair cell types are found in discrete regions of individual sensory, epithelia. The presence of multiple hair cell types in fishes provides strong support to the hypothesis of regional differences in the responses of individual otolithic sensory epithelia. The finding of hair cell types in fishes that closely resemble those found in amniote vestibular endorgans also suggests that hair cell heterogeneity arose earlier in the evolution of the vertebrate ear than previously thought.

HTB140 melanoma cells under proton irradiation and/or alkylating agents

NASA Astrophysics Data System (ADS)

Korićanac, L.; Petrović, I.; Privitera, G.; Cuttone, G.; Ristić-Fira, A.

2007-09-01

Chemoresistance is a major problem in the treatment of malignant melanoma. The mainstay of treatment for melanoma is the DNA-alkylating agent dacarbazine (DTIC). Fotemustine (FM), a member of the chloroethylnitrosourea group of alkylating agents, has also demonstrated significant antitumor effects in malignant melanoma. However, the intrinsic and acquired resistance of melanoma limits the clinical application of these drugs. Melanomas are also extremely radioresistant. With the objective of enhancing growth inhibition of melanoma cells, combined treatments of FM or DTIC with proton irradiation have been investigated. These effects were studied on HTB140 melanoma cell viability and proliferation. Cells exposed to treatment with FM and protons have shown inhibition of cell growth and significant reduction of proliferation capacity compared to single irradiation or drug treatment. Treatment with DTIC and protons has shown improved growth inhibition compared to appropriate single drug treatment, while the effects of single proton irradiation have been the most pronounced.

Transcriptome Analysis of Salicylic Acid Treatment in Rehmannia glutinosa Hairy Roots Using RNA-seq Technique for Identification of Genes Involved in Acteoside Biosynthesis

PubMed Central

Wang, Fengqing; Zhi, Jingyu; Zhang, Zhongyi; Wang, Lina; Suo, Yanfei; Xie, Caixia; Li, Mingjie; Zhang, Bao; Du, Jiafang; Gu, Li; Sun, Hongzheng

2017-01-01

Rehmannia glutinosa is a common bulk medicinal material that has been widely used in China due to its active ingredients. Acteoside, one of the ingredients, has antioxidant, antinephritic, anti-inflammatory, hepatoprotective, immunomodulatory, and neuroprotective effects, is usually selected as a quality-control component for R. glutinosa herb in the Chinese Pharmacopeia. The acteoside biosynthesis pathway in R. glutinosa has not yet been clearly established. Herein, we describe the establishment of a genetic transformation system for R. glutinosa mediated by Agrobacterium rhizogenes. We screened the optimal elicitors that markedly increased acteoside accumulation in R. glutinosa hairy roots. We found that acteoside accumulation dramatically increased with the addition of salicylic acid (SA); the optimal SA dose was 25 μmol/L for hairy roots. RNA-seq was applied to analyze the transcriptomic changes in hairy roots treated with SA for 24 h in comparison with an untreated control. A total of 3,716, 4,018, and 2,715 differentially expressed transcripts (DETs) were identified in 0 h-vs.-12 h, 0 h-vs.-24 h, and 12 h-vs.-24 h libraries, respectively. KEGG pathway-based analysis revealed that 127 DETs were enriched in “phenylpropanoid biosynthesis.” Of 219 putative unigenes involved in acteoside biosynthesis, 54 were found to be up-regulated at at least one of the time points after SA treatment. Selected candidate genes were analyzed by quantitative real-time PCR (qRT-PCR) in hairy roots with SA, methyl jasmonate (MeJA), AgNO3 (Ag+), and putrescine (Put) treatment. All genes investigated were up-regulated by SA treatment, and most candidate genes were weakly increased by MeJA to some degree. Furthermore, transcription abundance of eight candidate genes in tuberous roots of the high-acteoside-content (HA) cultivar QH were higher than those of the low-acteoside-content (LA) cultivar Wen 85-5. These results will pave the way for understanding the molecular basis of

Allelopathy in the natural and agricultural ecosystems and isolation of potent allelochemicals from Velvet bean (Mucuna pruriens) and Hairy vetch (Vicia villosa).

PubMed

Fujii, Yoshiharu

2003-06-01

We have studied on allelopathy of plants and developed methods to identify the effective substances in root exudates, leaf leacheate, and volatile chemicals emitted from plants. We found traditional cover plants that show allelopathic activity are useful for weed control. It could eliminate the use of synthetic chemicals for this purpose. Allelopathy is a natural power of plants to protect themselves by producing natural organic chemicals. Some endemic plants in Asia, already known by farmers in the region, as either cover crops used in intercropping, hedgerow, or agroforestry, were found to possess strong allelopathic abilities. Our group identified several allelochemicals from these plants. These allelopathic cover crops, mostly leguminous plants, provide protein rich food, and grow easily without artificial fertilizers, herbicides, insecticides and fungicides. In this regards, these allelopathic cover crops could save food shortage in rural area, and are useful for environmental conservation. Screenings of allelopathic plants by specific bioassays and field tests have been conducted. Hairy vetch (Vicia villosa) and Velvet bean (Mucuna pruriens) are two promising species for the practical application of allelopathy. An amino acid, L-DOPA, unusual in plants, plays an important role as allelochemical in Velvet bean (Mucuna pruriens). Hairy vetch is the most promising cover plant for the weed control in orchard, vegetable and rice production and even for landscape amendment in abandoned field in Japan. We have isolated "cyanamide", a well known nitrogen fertilizer, from Hairy vetch. This is the first finding of naturally produced cyanamide in the world.

Sensitivity of human dental pulp cells to eighteen chemical agents used for endodontic treatments in dentistry.

PubMed

Kobayashi, Morio; Tsutsui, Takeo W; Kobayashi, Tomoko; Ohno, Maki; Higo, Yukari; Inaba, Tomohiro; Tsutsui, Takeki

2013-01-01

To determine the adverse effects against human dental pulp tissue, the sensitivity of human dental pulp cells (D824 cells) to 18 chemical agents used for endodontic treatments in dentistry was examined. The cytotoxicity, as determined by a decrease in colony-forming ability of cells treated with the chemical agents, increased as the concentration increased. As a quantitative measure of the cytotoxic effect, LC(50), the concentration which induces a 50% lethality, was extrapolated from the concentration-response curves. The rank of the chemical agents according to their cytotoxic effect (LC(50)) was sodium arsenite > formaldehyde > hydrogen peroxide > zinc oxide > thymol ≈ iodoform ≈ eugenol > guaiacol > ethylenediaminetetraacetic acid ≈ iodine > procaine > lidocaine ≈ chloramphenicol ≈ m-cresol > calcium hydroxide ≈ sodium hypochlorite ≈ phenol ≈ p-phenolsulfonic acid. To compare the cytotoxicity and the levels of apoptosis and mRNA expression of five genes related to the function of dental pulp tissue, D824 cells treated with the LC(50) concentrations of chemical agents were assayed by the TUNEL method and quantitative reverse transcription polymerase chain reaction analysis, respectively. The inducibility of apoptotic cells and the level of mRNA expression of the genes varied with the chemical agents, indicating that both effects occurred independent of the rank of cytotoxic effect of the chemical agents. The results not only provide information concerning cytotoxicity of various chemical agents to human dental pulp cells, but also show an insight into the diversity of the pharmacodynamic action of the chemical agents.

Victorian spectacle: Julia Pastrana, the bearded and hairy female.

PubMed

Browne, Janet; Messenger, Sharon

2003-12-01

Julia Pastrana toured Europe in the late 1850s advertising herself as the 'Bearded and hairy Lady' or 'Nonedescript'. She suffered from a rare inherited disorder, not understood until the late 20th century, which manifested itself in facial distortion and considerable facial hair in the male pattern. Doctors, as well as sensation seekers, were very keen to examine her. Her story is unusual, not least because she was mummified after death by her husband-manager and continued to tour as a mounted exhibit for a number of decades. Indirectly, she participated in the evolutionary debate in Britain. In 1857, when she arrived in Britain from America, she was popularly known as the baboon-woman. When Darwin's Origin of Species was published, and evolutionary controversy about ape-ancestry was hot in the air, she was more often likened to the gorilla or orang-utan - as a possible specimen of a missing link.

Manipulating biological agents and cells in micro-scale volumes for applications in medicine

PubMed Central

Tasoglu, Savas; Gurkan, Umut Atakan; Wang, ShuQi

2013-01-01

Recent technological advances provide new tools to manipulate cells and biological agents in micro/nano-liter volumes. With precise control over small volumes, the cell microenvironment and other biological agents can be bioengineered; interactions between cells and external stimuli can be monitored; and the fundamental mechanisms such as cancer metastasis and stem cell differentiation can be elucidated. Technological advances based on the principles of electrical, magnetic, chemical, optical, acoustic, and mechanical forces lead to novel applications in point-of-care diagnostics, regenerative medicine, in vitro drug testing, cryopreservation, and cell isolation/purification. In this review, we first focus on the underlying mechanisms of emerging examples for cell manipulation in small volumes targeting applications such as tissue engineering. Then, we illustrate how these mechanisms impact the aforementioned biomedical applications, discuss the associated challenges, and provide perspectives for further development. PMID:23575660

Multi-Targeted Agents in Cancer Cell Chemosensitization: What We Learnt from Curcumin Thus Far.

PubMed

Bordoloi, Devivasha; Roy, Nand K; Monisha, Javadi; Padmavathi, Ganesan; Kunnumakkara, Ajaikumar B

2016-01-01

Research over the past several years has developed many mono-targeted therapies for the prevention and treatment of cancer, but it still remains one of the fatal diseases in the world killing 8.2 million people annually. It has been well-established that development of chemoresistance in cancer cells against mono-targeted chemotherapeutic agents by modulation of multiple survival pathways is the major cause of failure of cancer chemotherapy. Therefore, inhibition of these pathways by non-toxic multi-targeted agents may have profoundly high potential in preventing drug resistance and sensitizing cancer cells to chemotherapeutic agents. To study the potential of curcumin, a multi-targeted natural compound, obtained from the plant Turmeric (Curcuma longa) in combination with standard chemotherapeutic agents to inhibit drug resistance and sensitize cancer cells to these agents based on available literature and patents. An extensive literature survey was performed in PubMed and Google for the chemosensitizing potential of curcumin in different cancers published so far and the patents published during 2014-2015. Our search resulted in many in vitro, in vivo and clinical reports signifying the chemosensitizing potential of curcumin in diverse cancers. There were 160 in vitro studies, 62 in vivo studies and 5 clinical studies. Moreover, 11 studies reported on hybrid curcumin: the next generation of curcumin based therapeutics. Also, 34 patents on curcumin's biological activity have been retrieved. Altogether, the present study reveals the enormous potential of curcumin, a natural, non-toxic, multi-targeted agent in overcoming drug resistance in cancer cells and sensitizing them to chemotherapeutic drugs.

Chloroquine, an Endocytosis Blocking Agent, Inhibits Zika Virus Infection in Different Cell Models

PubMed Central

Delvecchio, Rodrigo; Higa, Luiza M.; Pezzuto, Paula; Valadão, Ana Luiza; Garcez, Patrícia P.; Monteiro, Fábio L.; Loiola, Erick C.; Dias, André A.; Silva, Fábio J. M.; Aliota, Matthew T.; Caine, Elizabeth A.; Osorio, Jorge E.; Bellio, Maria; O’Connor, David H.; Rehen, Stevens; de Aguiar, Renato Santana; Savarino, Andrea; Campanati, Loraine; Tanuri, Amilcar

2016-01-01

Zika virus (ZIKV) infection in utero might lead to microcephaly and other congenital defects. Since no specific therapy is available thus far, there is an urgent need for the discovery of agents capable of inhibiting its viral replication and deleterious effects. Chloroquine is widely used as an antimalarial drug, anti-inflammatory agent, and it also shows antiviral activity against several viruses. Here we show that chloroquine exhibits antiviral activity against ZIKV in Vero cells, human brain microvascular endothelial cells, human neural stem cells, and mouse neurospheres. We demonstrate that chloroquine reduces the number of ZIKV-infected cells in vitro, and inhibits virus production and cell death promoted by ZIKV infection without cytotoxic effects. In addition, chloroquine treatment partially reveres morphological changes induced by ZIKV infection in mouse neurospheres. PMID:27916837

Chloroquine, an Endocytosis Blocking Agent, Inhibits Zika Virus Infection in Different Cell Models.

PubMed

Delvecchio, Rodrigo; Higa, Luiza M; Pezzuto, Paula; Valadão, Ana Luiza; Garcez, Patrícia P; Monteiro, Fábio L; Loiola, Erick C; Dias, André A; Silva, Fábio J M; Aliota, Matthew T; Caine, Elizabeth A; Osorio, Jorge E; Bellio, Maria; O'Connor, David H; Rehen, Stevens; de Aguiar, Renato Santana; Savarino, Andrea; Campanati, Loraine; Tanuri, Amilcar

2016-11-29

Zika virus (ZIKV) infection in utero might lead to microcephaly and other congenital defects. Since no specific therapy is available thus far, there is an urgent need for the discovery of agents capable of inhibiting its viral replication and deleterious effects. Chloroquine is widely used as an antimalarial drug, anti-inflammatory agent, and it also shows antiviral activity against several viruses. Here we show that chloroquine exhibits antiviral activity against ZIKV in Vero cells, human brain microvascular endothelial cells, human neural stem cells, and mouse neurospheres. We demonstrate that chloroquine reduces the number of ZIKV-infected cells in vitro, and inhibits virus production and cell death promoted by ZIKV infection without cytotoxic effects. In addition, chloroquine treatment partially reveres morphological changes induced by ZIKV infection in mouse neurospheres.

Light-enabled reversible self-assembly and tunable optical properties of stable hairy nanoparticles

NASA Astrophysics Data System (ADS)

Chen, Yihuang; Wang, Zewei; He, Yanjie; Yoon, Young Jun; Jung, Jaehan; Zhang, Guangzhao; Lin, Zhiqun

2018-02-01

The ability to dynamically organize functional nanoparticles (NPs) via the use of environmental triggers (temperature, pH, light, or solvent polarity) opens up important perspectives for rapid and convenient construction of a rich variety of complex assemblies and materials with new structures and functionalities. Here, we report an unconventional strategy for crafting stable hairy NPs with light-enabled reversible and reliable self-assembly and tunable optical properties. Central to our strategy is to judiciously design amphiphilic star-like diblock copolymers comprising inner hydrophilic blocks and outer hydrophobic photoresponsive blocks as nanoreactors to direct the synthesis of monodisperse plasmonic NPs intimately and permanently capped with photoresponsive polymers. The size and shape of hairy NPs can be precisely tailored by modulating the length of inner hydrophilic block of star-like diblock copolymers. The perpetual anchoring of photoresponsive polymers on the NP surface renders the attractive feature of self-assembly and disassembly of NPs on demand using light of different wavelengths, as revealed by tunable surface plasmon resonance absorption of NPs and the reversible transformation of NPs between their dispersed and aggregated states. The dye encapsulation/release studies manifested that such photoresponsive NPs may be exploited as smart guest molecule nanocarriers. By extension, the star-like block copolymer strategy enables the crafting of a family of stable stimuli-responsive NPs (e.g., temperature- or pH-sensitive polymer-capped magnetic, ferroelectric, upconversion, or semiconducting NPs) and their assemblies for fundamental research in self-assembly and crystallization kinetics of NPs as well as potential applications in optics, optoelectronics, magnetic technologies, sensory materials and devices, catalysis, nanotechnology, and biotechnology.

Postnatal development of autonomic and sensory innervation of thoracic hairy skin in the rat. A histochemical, immunocytochemical, and radioenzymatic study.

PubMed

Schotzinger, R J; Landis, S C

1990-05-01

Histochemical, immunocytochemical, and radioenzymatic techniques were used to examine the neurotransmitter-related properties of the innervation of thoracic hairy skin in rats during adulthood and postnatal development. In the adult, catecholamine-containing fibers were associated with blood vessels and piloerector muscles, and ran in nerve bundles throughout the dermis. The distribution of tyrosine hydroxylase (TH)-immunoreactive (IR) fibers was identical. Neuronal fibers displaying neuropeptide Y (NPY) immunoreactivity were seen in association with blood vessels. Double-labeling studies suggested that most, if not all, NPY-IR fibers were also TH-IR and likewise most, if not all, vessel-associated TH-IR fibers were also NPY-IR. Calcitonin gene-related peptide (CGRP)-IR fibers were observed near and penetrating into the epidermis, in close association with hair follicles and blood vessels, and in nerve bundles. A similar distribution of substance P (SP)-IR fibers was evident. In adult animals treated as neonates with the sympathetic neurotoxin 6-hydroxydopamine, a virtual absence of TH-IR and NPY-IR fibers was observed, whereas the distribution of CGRP-IR and SP-IR fibers appeared unaltered. During postnatal development, a generalized increase in the number, fluorescence intensity, and varicose morphology of neuronal fibers displaying catecholamine fluorescence, NPY-IR, CGRP-IR, and SP-IR was observed. By postnatal day 21, the distribution of the above fibers had reached essentially adult levels, although the density of epidermal-associated CGRP-IR and SP-IR fibers was significantly greater than in the adult. The following were not evident in thoracic hairy skin at any timepoint examined: choline acetyltransferase activity, acetylcholinesterase histochemical staining or immunoreactivity, fibers displaying immunoreactivity to vasoactive intestinal peptide, cholecystokinin, or leucine-enkephalin. The present study demonstrates that the thoracic hairy skin in

Liposome encapsulation of chelating agents

DOEpatents

Rahman, Yueh Erh

1976-01-13

A method for transferring a chelating agent across a cellular membrane by encapsulating the charged chelating agent within liposomes and carrying the liposome-encapsulated chelating agent to the cellular membrane where the liposomes containing the chelating agent will be taken up by the cells, thereby transferring the chelating agent across the cellular membrane. A chelating agent can be introduced into the interior of a cell of a living organism wherein the liposomes will be decomposed, releasing the chelating agent to the interior of the cell. The released chelating agent will complex intracellularly deposited toxic heavy metals, permitting the more soluble metal complex to transfer across the cellular membrane from the cell and subsequently be removed from the living organism.

Effects of abscisic acid, gibberellin, ethylene and their interactions on production of phenolic acids in salvia miltiorrhiza bunge hairy roots.

PubMed

Liang, Zongsuo; Ma, Yini; Xu, Tao; Cui, Beimi; Liu, Yan; Guo, Zhixin; Yang, Dongfeng

2013-01-01

Salvia miltiorrhiza is one of the most important traditional Chinese medicinal plants because of its excellent performance in treating coronary heart disease. Phenolic acids mainly including caffeic acid, rosmarinic acid and salvianolic acid B are a group of active ingredients in S. miltiorrhiza. Abscisic acid (ABA), gibberellin (GA) and ethylene are three important phytohormones. In this study, effects of the three phytohormones and their interactions on phenolic production in S. miltiorrhiza hairy roots were investigated. The results showed that ABA, GA and ethylene were all effective to induce production of phenolic acids and increase activities of PAL and TAT in S. miltiorrhiza hairy roots. Effects of phytohormones were reversed by their biosynthetic inhibitors. Antagonistic actions between the three phytohormones played important roles in the biosynthesis of phenolic acids. GA signaling is necessary for ABA and ethylene-induced phenolic production. Yet, ABA and ethylene signaling is probably not necessary for GA3-induced phenolic production. The complex interactions of phytohormones help us reveal regulation mechanism of secondary metabolism and scale-up production of active ingredients in plants.

Effects of Abscisic Acid, Gibberellin, Ethylene and Their Interactions on Production of Phenolic Acids in Salvia miltiorrhiza Bunge Hairy Roots

PubMed Central

Xu, Tao; Cui, Beimi; Liu, Yan; Guo, Zhixin; Yang, Dongfeng

2013-01-01

Salvia miltiorrhiza is one of the most important traditional Chinese medicinal plants because of its excellent performance in treating coronary heart disease. Phenolic acids mainly including caffeic acid, rosmarinic acid and salvianolic acid B are a group of active ingredients in S. miltiorrhiza. Abscisic acid (ABA), gibberellin (GA) and ethylene are three important phytohormones. In this study, effects of the three phytohormones and their interactions on phenolic production in S. miltiorrhiza hairy roots were investigated. The results showed that ABA, GA and ethylene were all effective to induce production of phenolic acids and increase activities of PAL and TAT in S. miltiorrhiza hairy roots. Effects of phytohormones were reversed by their biosynthetic inhibitors. Antagonistic actions between the three phytohormones played important roles in the biosynthesis of phenolic acids. GA signaling is necessary for ABA and ethylene-induced phenolic production. Yet, ABA and ethylene signaling is probably not necessary for GA3-induced phenolic production. The complex interactions of phytohormones help us reveal regulation mechanism of secondary metabolism and scale-up production of active ingredients in plants. PMID:24023778

Matrix regeneration agents improve wound healing in non-stressed human corneal epithelial cells.

PubMed

Robciuc, A; Arvola, R P J; Jauhiainen, M; Holopainen, J M

2018-04-01

PurposeMatrix regenerating agents (RGTAs) emerged as promising in vivo wound-healing agents. These agents could prove beneficial for the treatment of dry eye disease-associated corneal micro-erosions; therefore, we aimed to evaluate the wound healing efficacy of regenerative agents (RGTAs or serum) in an in vitro model of hyperosmolarity (HO) stressed and non-stressed human corneal epithelial cells.Patients and methodsThe migration and proliferation induced by the regenerative agents was evaluated using an in vitro scratch wound assay and brome-deoxy-uridine incorporation. The inflammatory profile and effects of osmoregulators were also investigated. The two-tailed paired t-test calculated the statistical significance, with P-value<0.05 considered significant.ResultsThe most efficient inducer of re-epithelization was 2% serum, followed closely by 2% RGTA with an average improvement in cell migration of 1.8- and 1.4-fold, respectively, when compared with the non-treated control. Hyperosmolar stress significantly reduced the restorative effects of both serum and RGTAs; these effects were, however, neutralized by the osmoregulator betaine.ConclusionThese findings suggest that RGTAs could provide efficient treatment for dry-eye associated corneal micro-lesions if ocular surface HO is neutralized.

Sensitization of human carcinoma cells to alkylating agents by small interfering RNA suppression of 3-alkyladenine-DNA glycosylase.

PubMed

Paik, Johanna; Duncan, Tod; Lindahl, Tomas; Sedgwick, Barbara

2005-11-15

One of the major cytotoxic lesions generated by alkylating agents is DNA 3-alkyladenine, which can be excised by 3-alkyladenine DNA glycosylase (AAG). Inhibition of AAG may therefore result in increased cellular sensitivity to chemotherapeutic alkylating agents. To investigate this possibility, we have examined the role of AAG in protecting human tumor cells against such agents. Plasmids that express small interfering RNAs targeted to two different regions of AAG mRNA were transfected into HeLa cervical carcinoma cells and A2780-SCA ovarian carcinoma cells. Stable derivatives of both cell types with low AAG protein levels were sensitized to alkylating agents. Two HeLa cell lines with AAG protein levels reduced by at least 80% to 90% displayed a 5- to 10-fold increase in sensitivity to methyl methanesulfonate, N-methyl-N-nitrosourea, and the chemotherapeutic drugs temozolomide and 1,3-bis(2-chloroethyl)-1-nitrosourea. These cells showed no increase in sensitivity to UV light or ionizing radiation. After treatment with methyl methanesulfonate, AAG knockdown HeLa cells were delayed in S phase but accumulated in G2-M. Our data support the hypothesis that ablation of AAG activity in human tumor cells may provide a useful strategy to enhance the efficacy of current chemotherapeutic regimens that include alkylating agents.

Gum arabic glycoprotein is a twisted hairy rope. A new model based on O-galactosylhydroxyproline as the polysaccharide attachment site

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu Qi; Fong, C.; Lamport, D.T.A.

1991-07-01

Separation of the wound exudate from Acacia senegal (L.) Willd., gum arabic, on a preparative Superose-6 column gave two major fractions: a high molecular weight gum arabic glyco-protein (GAGP) containing about 90% carbohydrate and a lower molecular weight heterogeneous gum arabic polysaccharide fraction. Hydrogen fluoride-deglycosylation of GAGP gave a large hydroxyproline-rich polypeptide backbone (dGAGP). Alkaline hydrolysis of GAGP showed that most of the carbohydrate was attached to the polypeptide backbone as small hydroxyproline (Hyp)-polysaccharide substituents. The data imply a rodlike molecule with numerous small polysaccharide substituents (attached to 24% of the Hyp residues), regularly arranged along a highly periodic polypeptidemore » backbone based, hypothetically, on a 10 to 12 residue repetitive peptide motif. Thus, a simple statistical model of the gum arabic glycoprotein predicts a repeating polysaccharide substituents will maximize intramolecular hydrogen bonding if aligned along the long axis of the molecule, forming in effect a twisted hairy rope. Electron micrographs of rotary shadowed GAGP molecules support that prediction and may also explain show such apparently large molecules can exit the cell by endwise reptation through the small pores of the primary cell wall.« less

A novel anticancer agent SNG1153 inhibits growth of lung cancer stem/progenitor cells

PubMed Central

Wang, Jing; Zhu, Hai; Han, Yuqing; Jin, Mingji; Wang, Jun; Zhou, Congya; Ma, Junfeng; Lin, Qingcong; Wang, Zhaoyi; Meng, Kun; Fu, Xueqi

2016-01-01

Lung cancer is the leading cause of cancer-related death in both men and women. Lung cancer contains a small population of cancer cells with stem-like features known as cancer stem cells (CSCs). CSCs are often more resistant to current therapeutic treatments. Thus, it is urgent to develop a novel agent that is able to inhibit CSCs growth. In this study, we examined the ability of SNG1153, a novel chemical agent to inhibit the growth of lung CSCs. We found that SNG1153 inhibited growth and induced apoptosis in established lung cancer cells. We also found that SNG1153 inhibited the tumorsphere formation and decreased CD133-positive (lung CSC marker) cancer cells. SNG1153 was able to attenuate tumor formation in NOD/SCID (non-obese diabetic/severe combined immunodeficient) mice injected with lung tumorsphere cells. We further demonstrated that SNG1153 induced β-catenin phosphorylation and down-regulated β-catenin. Our results thus demonstrate that SNG1153 effectively inhibits the growth of lung CSCs and suggest that SNG1153 may be a novel therapeutic agent to treat human lung cancer. PMID:27281614

Insect adhesion on rough surfaces: analysis of adhesive contact of smooth and hairy pads on transparent microstructured substrates

PubMed Central

Zhou, Yanmin; Robinson, Adam; Steiner, Ullrich; Federle, Walter

2014-01-01

Insect climbing footpads are able to adhere to rough surfaces, but the details of this capability are still unclear. To overcome experimental limitations of randomly rough, opaque surfaces, we fabricated transparent test substrates containing square arrays of 1.4 µm diameter pillars, with variable height (0.5 and 1.4 µm) and spacing (from 3 to 22 µm). Smooth pads of cockroaches (Nauphoeta cinerea) made partial contact (limited to the tops of the structures) for the two densest arrays of tall pillars, but full contact (touching the substrate in between pillars) for larger spacings. The transition from partial to full contact was accompanied by a sharp increase in shear forces. Tests on hairy pads of dock beetles (Gastrophysa viridula) showed that setae adhered between pillars for larger spacings, but pads were equally unable to make full contact on the densest arrays. The beetles' shear forces similarly decreased for denser arrays, but also for short pillars and with a more gradual transition. These observations can be explained by simple contact models derived for soft uniform materials (smooth pads) or thin flat plates (hairy-pad spatulae). Our results show that microstructured substrates are powerful tools to reveal adaptations of natural adhesives for rough surfaces. PMID:24990289

Insect adhesion on rough surfaces: analysis of adhesive contact of smooth and hairy pads on transparent microstructured substrates.

PubMed

Zhou, Yanmin; Robinson, Adam; Steiner, Ullrich; Federle, Walter

2014-09-06

Insect climbing footpads are able to adhere to rough surfaces, but the details of this capability are still unclear. To overcome experimental limitations of randomly rough, opaque surfaces, we fabricated transparent test substrates containing square arrays of 1.4 µm diameter pillars, with variable height (0.5 and 1.4 µm) and spacing (from 3 to 22 µm). Smooth pads of cockroaches (Nauphoeta cinerea) made partial contact (limited to the tops of the structures) for the two densest arrays of tall pillars, but full contact (touching the substrate in between pillars) for larger spacings. The transition from partial to full contact was accompanied by a sharp increase in shear forces. Tests on hairy pads of dock beetles (Gastrophysa viridula) showed that setae adhered between pillars for larger spacings, but pads were equally unable to make full contact on the densest arrays. The beetles' shear forces similarly decreased for denser arrays, but also for short pillars and with a more gradual transition. These observations can be explained by simple contact models derived for soft uniform materials (smooth pads) or thin flat plates (hairy-pad spatulae). Our results show that microstructured substrates are powerful tools to reveal adaptations of natural adhesives for rough surfaces.

Comparison of the clinical efficacy between single-agent and dual-agent concurrent chemoradiotherapy in the treatment of unresectable esophageal squamous cell carcinoma: a multicenter retrospective analysis.

PubMed

Li, Jie; Gong, Youling; Diao, Peng; Huang, Qingmei; Wen, Yixue; Lin, Binwei; Cai, Hongwei; Tian, Honggang; He, Bing; Ji, Lanlan; Guo, Ping; Miao, Jidong; Du, Xiaobo

2018-01-22

Some Chinese patients with esophageal squamous cell carcinomaare often treated with single-agent concurrent chemoradiotherapy. However, no results have been reported from randomized controlled clinical trials comparing single-agent with double-agent concurrent chemoradiotherapy. It therefore remains unclear whether these regimens are equally clinically effective. In this study, we retrospectively analyzed and compared the therapeutic effects of single-agent and double-agent concurrent chemoradiotherapy in patients with unresectable esophageal squamous cell carcinoma. This study enrolled 168 patients who received definitive concurrent chemoradiotherapy for locally advanced unresectable esophageal squamous carcinoma at 10 hospitals between 2010 and 2015. We evaluated survival time and toxicity. The Kaplan-Meier method was used to estimate survival data. The log-rank test was used in univariate analysis A Cox proportional hazards regression model was used to conduct a multivariate analysis of the effects of prognostic factors on survival. In this study, 100 (59.5%) and 68 patients (40.5%) received single-agent and dual-agent combination chemoradiotherapy, respectively. The estimate 5-year progression-free survival (PFS) rate and overall survival (OS) rate of dual-agent therapy was higher than that of single-agent therapy (52.5% and 40.9%, 78.2% and 60.7%, respectively), but there were no significant differences (P = 0.367 and 0.161, respectively). Multivariate analysis showed that sex, age,and radiotherapy dose had no significant effects on OS or PFS. Only disease stage was associated with OS and PFS in the multivariable analysis (P = 0.006 and 0.003, respectively). In dual-agent group, the incidence of acute toxicity and the incidence of 3 and4 grade toxicity were higher than single-agent group. The 5-year PFS and OS rates of dual-agent therapy were higher than those of single-agent concurrent chemoradiotherapy for patients with unresectable esophageal

Water as foaming agent for open cell polyurethane structures.

PubMed

Haugen, H; Ried, V; Brunner, M; Will, J; Wintermantel, E

2004-04-01

The problem of moisture in polymer processing is known to any polymer engineer, as air bubbles may be formed. Hence granulates are generally dried prior to manufacturing. This study tried to develop a novel processing methods for scaffolds with controlled moisture content in thermoplastic polyurethane. The common foaming agents for polyurethane are organic solvents, whose residues remaining in the scaffold may be harmful to adherent cells, protein growth factors or nearby tissues. Water was used as a foaming agent and NaCl was used as porogens to achieve an open-cell structure. The polyether-polyurethane samples were processed in a heated press, and achieved a porosity of 64%. The pore size ranged between 50 and 500 microm. Human fibroblasts adhered and proliferate in the scaffold. A non-toxic production process was developed to manufacture a porous structure with a thermoplastic polyether-polyurethane. The process enables a mass-production of samples with adjustable pore size and porosity. In contrast to an existing method (solvent casting), the processing of the samples was not limited by its thickness. The process parameters, which attribute mostly to the pore building, were filling volume, temperature, NaCl-concentration and water-uptake rate.

A quantitative risk assessment of exposure to adventitious agents in a cell culture-derived subunit influenza vaccine.

PubMed

Gregersen, Jens-Peter

2008-06-19

A risk-assessment model has demonstrated the ability of a new cell culture-based vaccine manufacturing process to reduce the level of any adventitious agent to a million-fold below infectious levels. The cell culture-derived subunit influenza vaccine (OPTAFLU), Novartis Vaccines and Diagnostics) is produced using Madin-Darby canine kidney (MDCK) cells to propagate seasonal viral strains, as an alternative to embryonated chicken-eggs. As only a limited range of mammalian viruses can grow in MDCK cells, similar to embryonated eggs, MDCK cells can act as an effective filter for a wide range of adventitious agents that might be introduced during vaccine production. However, the introduction of an alternative cell substrate (for example, MDCK cells) into a vaccine manufacturing process requires thorough investigations to assess the potential for adventitious agent risk in the final product, in the unlikely event that contamination should occur. The risk assessment takes into account the entire manufacturing process, from initial influenza virus isolation, through to blending of the trivalent subunit vaccine and worst-case residual titres for the final vaccine formulation have been calculated for >20 viruses or virus families. Maximum residual titres for all viruses tested were in the range of 10(-6) to 10(-16) infectious units per vaccine dose. Thus, the new cell culture-based vaccine manufacturing process can reduce any adventitious agent to a level that is unable to cause infection.

Tolerance of dormant and active cells in Pseudomonas aeruginosa PA01 biofilm to antimicrobial agents.

PubMed

Kim, Jaeeun; Hahn, Ji-Sook; Franklin, Michael J; Stewart, Philip S; Yoon, Jeyong

2009-01-01

The aim of the study was to determine the susceptibility of active and dormant cell populations from Pseudomonas aeruginosa biofilms to non-antibiotic antimicrobial agents such as chlorine, hydrogen peroxide and silver ions in comparison with antibiotics. Active cells in colony biofilm were differentially labelled by induction of a green fluorescent protein (GFP). Active and dormant cells were sorted in phosphate buffered solution by flow cytometry. Reductions in viability were determined with plate counts. The spatial pattern of metabolic activity in colony biofilm was verified, and the active and dormant cells were successfully sorted according to the GFP intensity. Active cells had bigger cell size and higher intracellular density than dormant cells. While dormant cells were more tolerant to tobramycin and silver ions, active cells were more tolerant to chlorine. Metabolically active cells contain denser intracellular components that can react with highly reactive oxidants such as chlorine, thereby reducing the available concentrations of chlorine. In contrast, the concentrations of silver ions and hydrogen peroxide were constant during treatment. Aerobically grown stationary cells were significantly more tolerant to chlorine unlike other antimicrobial agents. Chlorine was more effective in inactivation of metabolically inactive dormant cells and also more effective under anaerobic conditions. The high oxidative reactivity and rapid decay of chlorine might influence the different antimicrobial actions of chlorine compared with antibiotics. This study contributes to understanding the effects of dormancy and the presence of oxygen on the susceptibility of P. aeruginosa biofilm to a wide range of antimicrobial agents.

Evaluation of tobacco (Nicotiana tabacum L. cv. Petit Havana SR1) hairy roots for the production of geraniol, the first committed step in terpenoid indole alkaloid pathway.

PubMed

Ritala, Anneli; Dong, Lemeng; Imseng, Nicole; Seppänen-Laakso, Tuulikki; Vasilev, Nikolay; van der Krol, Sander; Rischer, Heiko; Maaheimo, Hannu; Virkki, Arho; Brändli, Johanna; Schillberg, Stefan; Eibl, Regine; Bouwmeester, Harro; Oksman-Caldentey, Kirsi-Marja

2014-04-20

The terpenoid indole alkaloids are one of the major classes of plant-derived natural products and are well known for their many applications in the pharmaceutical, fragrance and cosmetics industries. Hairy root cultures are useful for the production of plant secondary metabolites because of their genetic and biochemical stability and their rapid growth in hormone-free media. Tobacco (Nicotiana tabacum L. cv. Petit Havana SR1) hairy roots, which do not produce geraniol naturally, were engineered to express a plastid-targeted geraniol synthase gene originally isolated from Valeriana officinalis L. (VoGES). A SPME-GC-MS screening tool was developed for the rapid evaluation of production clones. The GC-MS analysis revealed that the free geraniol content in 20 hairy root clones expressing VoGES was an average of 13.7 μg/g dry weight (DW) and a maximum of 31.3 μg/g DW. More detailed metabolic analysis revealed that geraniol derivatives were present in six major glycoside forms, namely the hexose and/or pentose conjugates of geraniol and hydroxygeraniol, resulting in total geraniol levels of up to 204.3 μg/g DW following deglycosylation. A benchtop-scale process was developed in a 20-L wave-mixed bioreactor eventually yielding hundreds of grams of biomass and milligram quantities of geraniol per cultivation bag. Copyright © 2014 Elsevier B.V. All rights reserved.

Induced Biosynthesis of resveratrol and the prenylated stilbenoids arachidan-1 and arachidan-3 in hairy root cultures of peanut: effects of culture medium and growth stage

USDA-ARS?s Scientific Manuscript database

The peanut plant has evolved specialized biosynthetic mechanisms that allowed resisting infection by producing diverse secondary metabolites. Among these unique compounds are the stilbenoids, which include resveratrol analogues. Our previous research demonstrated that peanut hairy root cultures prov...

Inhibition of the Nedd8 system sensitizes cells to DNA Inter-strand crosslinking agents

PubMed Central

Kee, Younghoon; Huang, Min; Chang, Sophia; Moreau, Lisa A.; Park, Eunmi; Smith, Peter G.; D’Andrea, Alan D.

2012-01-01

The Fanconi Anemia (FA) pathway is required for repair of DNA interstrand crosslinks (ICLs). FA pathway-deficient cells are hypersensitive to DNA ICL-inducing drugs such as Cisplatin. Conversely, hyperactivation of the FA pathway is a mechanism that may underlie cellular resistance to DNA ICL agents. Modulating FANCD2 monoubiquitination, a key step in the FA pathway, may be an effective therapeutic approach to conferring cellular sensitivity to ICL agents. Here, we show that inhibition of the Nedd8 conjugation system increases cellular sensitivity to DNA ICL-inducing agents. Mechanistically, the Nedd8 inhibition, either by siRNA-mediated knockdown of Nedd8 conjugating enzymes or treatment with a Nedd8 activating enzyme inhibitor MLN4924, suppressed DNA damage-induced FANCD2 monoubiquitination and CHK1 phosphorylation. Our data indicate that inhibition of the FA pathway is largely responsible for the heightened cellular sensitivity to DNA ICLs upon Nedd8 inhibition. These results suggest that a combination of Nedd8 inhibition with ICL-inducing agents may be an effective strategy for sensitizing a subset of drug-resistant cancer cells. PMID:22219386

Nitric oxide donors attenuate clongenic potential in rat C6 glioma cells treated with alkylating chemotherapeutic agents.

PubMed

Yang, Jir-Jei; Yin, Jiu-Haw; Yang, Ding-I

2007-05-11

1,3-Bis(2-chloroethyl)-1-nitrosourea (BCNU) kills tumor cells via multiple actions including alkylation and carbamoylation. Previously, we have reported that formation of S-nitrosoglutathione (GSNO) in glioma cells overexpressing inducible nitric oxide synthase (iNOS) contributed to nitric oxide (NO)-dependent carbamoylating chemoresistance against BCNU. To further characterize the effects of NO on alkylating cytotoxicity, colony formation assay was applied to evaluate the effects of various NO donors on rat C6 glioma cells challenged with alkylating agents. We demonstrate that NO donors including GSNO, diethylamine NONOate (DEA/NO), and sodium nitroprusside (SNP) substantially reduced the extent of colony formation in glioma cells treated with alkylating agents, namely methyl methanesulfonate (MMS), N-methyl-N-nitrosourea (MNU), and N-ethyl-N-nitrosourea (ENU). Without alkylating agents these NO-releasing agents alone had no effects on clongenic potential of rat C6 glioma cells. Among these three NO donors used, the effectiveness in potentiating alkylating cytotoxicity is in the order of "GSNO>DEA/NO>SNP" when applied at the same dosages. GSNO also exerted similar synergistic actions reducing the extents of colony formation when co-administrated with 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-hydrazine (compound #1), another alkylating agent that mimics the chloroethylating action of BCNU. Together with our previous findings, we propose that NO donors may be used as adjunct chemotherapy with alkylating agents for such malignant brain tumors as glioblastoma multiforme (GBM). In contrast, production of NO as a result of iNOS induction, such as that occurring after surgical resection of brain tumors, may compromise the efficacy of carbamoylating chemotherapy.

Alternative oxidase (AOX) and phenolic metabolism in methyl jasmonate-treated hairy root cultures of Daucus carota L.

PubMed

Sircar, Debabrata; Cardoso, Hélia G; Mukherjee, Chiranjit; Mitra, Adinpunya; Arnholdt-Schmitt, Birgit

2012-05-01

Methyl-jasmonate (MJ)-treated hairy roots of Daucus carota L. were used to study the influence of alternative oxidase (AOX) in phenylpropanoid metabolism. Phenolic acid accumulation, as well as total flavonoids and lignin content of the MJ-treated hairy roots were decreased by treatment with salicylhydroxamic acid (SHAM), a known inhibitor of AOX. The inhibitory effect of SHAM was concentration dependent. Treatment with propyl gallate (PG), another inhibitor of AOX, also had a similar inhibitory effect on accumulation of phenolic acid, total flavonoids and lignin. The transcript levels of two DcAOX genes (DcAOX2a and DcAOX1a) were monitored at selected post-elicitation time points. A notable rise in the transcript levels of both DcAOX genes was observed preceding the MJ-induced enhanced accumulation of phenolics, flavonoids and lignin. An appreciable increase in phenylalanine ammonia-lyase (PAL) transcript level was also observed prior to enhanced phenolics accumulation. Both DcAOX genes showed differential transcript accumulation patterns after the onset of elicitation. The transcript levels of DcAOX1a and DcAOX2a attained peak at 6hours post elicitation (hpe) and 12hpe, respectively. An increase in the transcript levels of both DcAOX genes preceding the accumulation of phenylpropanoid-derivatives and lignin showed a positive correlation between AOX activity and phenylpropanoid biosynthesis. The results provide important new insight about the influence of AOX in phenylpropanoid biosynthesis. Copyright © 2012 Elsevier GmbH. All rights reserved.

Strategies to overcome oxygen transfer limitations during hairy root cultivation of Azadiracta indica for enhanced azadirachtin production.

PubMed

Srivastava, Smita; Srivastava, Ashok Kumar

2012-07-01

The vast untapped potential of hairy root cultures as a stable source of biologically active chemicals has focused the attention of scientific community toward its commercial exploitation. However, the major bottleneck remains its successful scale-up. Due to branching, the roots form an interlocked matrix that exhibits resistance to oxygen transfer. Thus, present work was undertaken to develop cultivation strategies like optimization of inlet gas composition (in terms of % (v/v) O(2) in air), air-flow rate and addition of oxygen vectors in the medium, to curb the oxygen transfer limitations during hairy root cultivation of Azadirachta indica for in vitro azadirachtin (a biopesticide) production. It was found that increasing the oxygen fraction in the inlet air (in the range, 20-100% (v/v) O(2) in air) increased the azadirachtin productivity by approximately threefold, to a maximum of 4.42 mg/L per day (at 100% (v/v) O(2) in air) with respect to 1.68 mg/L per day in control (air with no oxygen supplementation). Similarly, increasing the air-flow rate (in the range, 0.3-2 vvm) also increased the azadirachtin productivity to a maximum of 1.84 mg/L per day at 0.8 vvm of air-flow rate. On the contrary, addition of oxygen vectors (in the range, 1-4% (v/v); hydrogen peroxide, toluene, Tween 80, kerosene, silicone oil, and n-hexadecane), decreased the azadirachtin productivity with respect to control (1.76 mg/L per day).

Biosynthesis of luminescent CdS quantum dots using plant hairy root culture

NASA Astrophysics Data System (ADS)

Borovaya, Mariya N.; Naumenko, Antonina P.; Matvieieva, Nadia A.; Blume, Yaroslav B.; Yemets, Alla I.

2014-12-01

CdS nanoparticles have a great potential for application in chemical research, bioscience and medicine. The aim of this study was to develop an efficient and environmentally-friendly method of plant-based biosynthesis of CdS quantum dots using hairy root culture of Linaria maroccana L. By incubating Linaria root extract with inorganic cadmium sulfate and sodium sulfide we synthesized stable luminescent CdS nanocrystals with absorption peaks for UV-visible spectrometry at 362 nm, 398 nm and 464 nm, and luminescent peaks at 425, 462, 500 nm. Transmission electron microscopy of produced quantum dots revealed their spherical shape with a size predominantly from 5 to 7 nm. Electron diffraction pattern confirmed the wurtzite crystalline structure of synthesized cadmium sulfide quantum dots. These results describe the first successful attempt of quantum dots synthesis using plant extract.

Genetically Modified T-cell Infusion Following Peripheral Blood Stem Cell Transplant in Treating Patients With Recurrent or High-Risk Non-Hodgkin Lymphoma

ClinicalTrials.gov

2018-01-26

Adult Grade III Lymphomatoid Granulomatosis; Cutaneous B-cell Non-Hodgkin Lymphoma; Extranodal Marginal Zone B-cell Lymphoma of Mucosa-associated Lymphoid Tissue; Intraocular Lymphoma; Nodal Marginal Zone B-cell Lymphoma; Post-transplant Lymphoproliferative Disorder; Recurrent Adult Burkitt Lymphoma; Recurrent Adult Diffuse Large Cell Lymphoma; Recurrent Adult Diffuse Mixed Cell Lymphoma; Recurrent Adult Diffuse Small Cleaved Cell Lymphoma; Recurrent Adult Grade III Lymphomatoid Granulomatosis; Recurrent Adult Immunoblastic Large Cell Lymphoma; Recurrent Adult Lymphoblastic Lymphoma; Recurrent Grade 1 Follicular Lymphoma; Recurrent Grade 2 Follicular Lymphoma; Recurrent Grade 3 Follicular Lymphoma; Recurrent Mantle Cell Lymphoma; Recurrent Marginal Zone Lymphoma; Recurrent Small Lymphocytic Lymphoma; Refractory Hairy Cell Leukemia; Small Intestine Lymphoma; Splenic Marginal Zone Lymphoma; Testicular Lymphoma; Waldenström Macroglobulinemia

Transgenic analysis reveals LeACS-1 as a positive regulator of ethylene-induced shikonin biosynthesis in Lithospermum erythrorhizon hairy roots.

PubMed

Fang, Rongjun; Wu, Fengyao; Zou, Ailan; Zhu, Yu; Zhao, Hua; Zhao, Hu; Liao, Yonghui; Tang, Ren-Jie; Yang, Tongyi; Pang, Yanjun; Wang, Xiaoming; Yang, Rongwu; Qi, Jinliang; Lu, Guihua; Yang, Yonghua

2016-03-01

The phytohormone ethylene (ET) is a crucial signaling molecule that induces the biosynthesis of shikonin and its derivatives in Lithospermum erythrorhizon shoot cultures. However, the molecular mechanism and the positive regulators involved in this physiological process are largely unknown. In this study, the function of LeACS-1, a key gene encoding the 1-aminocyclopropane-1-carboxylic acid synthase for ET biosynthesis in L. erythrorhizon hairy roots, was characterized by using overexpression and RNA interference (RNAi) strategies. The results showed that overexpression of LeACS-1 significantly increased endogenous ET concentration and shikonin production, consistent with the up-regulated genes involved in ET biosynthesis and transduction, as well as the genes related to shikonin biosynthesis. Conversely, RNAi of LeACS-1 effectively decreased endogenous ET concentration and shikonin production and down-regulated the expression level of above genes. Correlation analysis showed a significant positive linear relationship between ET concentration and shikonin production. All these results suggest that LeACS-1 acts as a positive regulator of ethylene-induced shikonin biosynthesis in L. erythrorhizon hairy roots. Our work not only gives new insights into the understanding of the relationship between ET and shikonin biosynthesis, but also provides an efficient genetic engineering target gene for secondary metabolite production in non-model plant L. erythrorhizon.

Effect of Medium Supplements on Agrobacterium rhizogenes Mediated Hairy Root Induction from the Callus Tissues of Camellia sinensis var. sinensis

PubMed Central

Rana, Mohammad M.; Han, Zhuo-Xiao; Song, Da-Peng; Liu, Guo-Feng; Li, Da-Xiang; Wan, Xiao-Chun; Karthikeyan, Alagarsamy; Wei, Shu

2016-01-01

Tea (Camellia sinensis L.) is recalcitrant to Agrobacterium-mediated genetic transformation largely due to the bactericidal effects of tea polyphenols and phenolics oxidation induced by necrosis of explant tissue over the process of transformation. In this study, different antioxidants/adsorbents were added as supplements to the co-cultivation and post co-cultivation media to overcome these problems for the transformation improvement. Tea-cotyledon-derived calli were used as explants and Agrobacterium rhizognes strain ATCC 15834 was used as a mediator. Results showed that Agrobacterium growth, virulence (vir) gene expression and browning of explant tissue were greatly influenced by different supplements. Murashige and Skoog (MS) basal salts medium supplemented with 30 g·L−1 sucrose, 0.1 g·L−1 l-glutamine and 5 g·L−1 polyvinylpolypyrrolidone (PVPP) as co-cultivation and post co-cultivation media could maintain these parameters better that ultimately led to significant improvement of hairy root generation efficiency compared to that in the control (MS + 30 g·L−1 sucrose). Additionally, the reporter genes β-glucuronidase (gusA) and cyan fluorescent protein (cfp) were also stably expressed in the transgenic hairy roots. Our study would be helpful in establishing a feasible approach for tea biological studies and genetic improvement of tea varieties. PMID:27428960

Effect of Medium Supplements on Agrobacterium rhizogenes Mediated Hairy Root Induction from the Callus Tissues of Camellia sinensis var. sinensis.

PubMed

Rana, Mohammad M; Han, Zhuo-Xiao; Song, Da-Peng; Liu, Guo-Feng; Li, Da-Xiang; Wan, Xiao-Chun; Karthikeyan, Alagarsamy; Wei, Shu

2016-07-15

Tea (Camellia sinensis L.) is recalcitrant to Agrobacterium-mediated genetic transformation largely due to the bactericidal effects of tea polyphenols and phenolics oxidation induced by necrosis of explant tissue over the process of transformation. In this study, different antioxidants/adsorbents were added as supplements to the co-cultivation and post co-cultivation media to overcome these problems for the transformation improvement. Tea-cotyledon-derived calli were used as explants and Agrobacterium rhizognes strain ATCC 15834 was used as a mediator. Results showed that Agrobacterium growth, virulence (vir) gene expression and browning of explant tissue were greatly influenced by different supplements. Murashige and Skoog (MS) basal salts medium supplemented with 30 g·L(-1) sucrose, 0.1 g·L(-1) l-glutamine and 5 g·L(-1) polyvinylpolypyrrolidone (PVPP) as co-cultivation and post co-cultivation media could maintain these parameters better that ultimately led to significant improvement of hairy root generation efficiency compared to that in the control (MS + 30 g·L(-1) sucrose). Additionally, the reporter genes β-glucuronidase (gusA) and cyan fluorescent protein (cfp) were also stably expressed in the transgenic hairy roots. Our study would be helpful in establishing a feasible approach for tea biological studies and genetic improvement of tea varieties.

A Novel Isoquinoline Derivative Anticancer Agent and Its Targeted Delivery to Tumor Cells Using Transferrin-Conjugated Liposomes

PubMed Central

Yang, Xuewei; Yang, Shuang; Chai, Hongyu; Yang, Zhaogang; Lee, Robert J.; Liao, Weiwei; Teng, Lesheng

2015-01-01

We have screened 11 isoquinoline derivatives and α-methylene-γ-butyrolactones using the 3-(4,5-dimethylthi-azol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity assay in HeLa and HEK-293T cells. Compound 2 was identified as potential anticancer agent. To further improve its therapeutic potential, this agent was incorporated into transferrin (Tf)-conjugated liposomes (LPs) for targeted delivery to tumor cells. We have demonstrated Tf-LP-Compound 2 have superior antitumor activity compared to non-targeted controls and the free drug. These data show Tf-LP-Compound 2 to be a promising agent that warrants further evaluation. PMID:26309138

Chemically synthesized silver nanoparticles as cell lysis agent for bacterial genomic DNA isolation

NASA Astrophysics Data System (ADS)

Goswami, Gunajit; Boruah, Himangshu; Gautom, Trishnamoni; Jyoti Hazarika, Dibya; Barooah, Madhumita; Boro, Robin Chandra

2017-12-01

Silver nanoparticles (AgNPs) have seen a recent spurt of use in varied fields of science. In this paper, we showed a novel application of AgNP as a promising microbial cell-lysis agent for genomic DNA isolation. We utilized chemically synthesized AgNPs for lysing bacterial cells to isolate their genomic DNA. The AgNPs efficiently lysed bacterial cells to yield good quality DNA that could be subsequently used for several molecular biology works.

Increased sensitivity of p53-deficient cells to anticancer agents due to loss of Pms2

PubMed Central

Fedier, A; Ruefenacht, U B; Schwarz, V A; Haller, U; Fink, D

2002-01-01

A large fraction of human tumours carries mutations in the p53 gene. p53 plays a central role in controlling cell cycle checkpoint regulation, DNA repair, transcription, and apoptosis upon genotoxic stress. Lack of p53 function impairs these cellular processes, and this may be the basis of resistance to chemotherapeutic regimens. By virtue of the involvement of DNA mismatch repair in modulating cytotoxic pathways in response to DNA damaging agents, we investigated the effects of loss of Pms2 on the sensitivity to a panel of widely used anticancer agents in E1A/Ha-Ras-transformed p53-null mouse fibroblasts either proficient or deficient in Pms2. We report that lack of the Pms2 gene is associated with an increased sensitivity, ranging from 2–6-fold, to some types of anticancer agents including the topoisomerase II poisons doxorubicin, etoposide and mitoxantrone, the platinum compounds cisplatin and oxaliplatin, the taxanes docetaxel and paclitaxel, and the antimetabolite gemcitabine. In contrast, no change in sensitivity was found after treatment with 5-fluorouracil. Cell cycle analysis revealed that both, Pms2-deficient and -proficient cells, retain the ability to arrest at the G2/M upon cisplatin treatment. The data indicate that the concomitant loss of Pms2 function chemosensitises p53-deficient cells to some types of anticancer agents, that Pms2 positively modulates cell survival by mechanisms independent of p53, and that increased cytotoxicity is paralleled by increased apoptosis. Tumour-targeted functional inhibition of Pms2 may be a valuable strategy for increasing the efficacy of anticancer agents in the treatment of p53-mutant cancers. British Journal of Cancer (2002) 87, 1027–1033. doi:10.1038/sj.bjc.6600599 www.bjcancer.com © 2002 Cancer Research UK PMID:12434296

Cell-permeable Ln(III) chelate-functionalized InP quantum dots as multimodal imaging agents.

PubMed

Stasiuk, Graeme J; Tamang, Sudarsan; Imbert, Daniel; Poillot, Cathy; Giardiello, Marco; Tisseyre, Céline; Barbier, Emmanuel L; Fries, Pascal Henry; de Waard, Michel; Reiss, Peter; Mazzanti, Marinella

2011-10-25

Quantum dots (QDs) are ideal scaffolds for the development of multimodal imaging agents, but their application in clinical diagnostics is limited by the toxicity of classical CdSe QDs. A new bimodal MRI/optical nanosized contrast agent with high gadolinium payload has been prepared through direct covalent attachment of up to 80 Gd(III) chelates on fluorescent nontoxic InP/ZnS QDs. It shows a high relaxivity of 900 mM(-1) s(-1) (13 mM(-1 )s(-1) per Gd ion) at 35 MHz (0.81 T) and 298 K, while the bright luminescence of the QDs is preserved. Eu(III) and Tb(III) chelates were also successfully grafted to the InP/ZnS QDs. The absence of energy transfer between the QD and lanthanide emitting centers results in a multicolor system. Using this convenient direct grafting strategy additional targeting ligands can be included on the QD. Here a cell-penetrating peptide has been co-grafted in a one-pot reaction to afford a cell-permeable multimodal multimeric MRI contrast agent that reports cellular localization by fluorescence and provides high relaxivity and increased tissue retention with respect to commercial contrast agents.

Suspected pyrrolizidine alkaloid hepatotoxicosis in wild southern hairy-nosed wombats (Lasiorhinus latifrons).

PubMed

Woolford, Lucy; Fletcher, Mary T; Boardman, Wayne S J

2014-07-30

Southern hairy-nosed wombats (Lasiorhinus latifrons) inhabiting degraded habitat in South Australia were recently identified with extensive hair loss and dermatitis and were in thin to emaciated body condition. Pathological and clinicopathological investigations on affected juvenile wombats identified a toxic hepatopathy suggestive of plants containing pyrrolizidine alkaloids, accompanied by photosensitive dermatitis. Hepatic disease was suspected in additional wombats on the basis of serum biochemical analysis. Preliminary toxicological analysis performed on scats and gastrointestinal contents from wombats found in this degraded habitat identified a number of toxic pyrrolizidine alkaloids consistent with ingestion of Heliotropeum europaeum. Although unpalatable, ingestion may occur by young animals due to decreased availability of preferred forages in degraded habitats and the emergence of weeds around the time of weaning of naive animals. Habitat degradation leading to malnutrition and ingestion of toxic weed species is a significant welfare issue in this species.

Hairy root transgene expression analysis of a secretory peroxidase (PvPOX1) from common bean infected by Fusarium wilt.

PubMed

Xue, Renfeng; Wu, Xingbo; Wang, Yingjie; Zhuang, Yan; Chen, Jian; Wu, Jing; Ge, Weide; Wang, Lanfen; Wang, Shumin; Blair, Matthew W

2017-07-01

Plant peroxidases (POXs) are one of the most important redox enzymes in the defense responses. However, the large number of different plant POX genes makes it necessary to carefully confirm the function of each paralogous POX gene in specific tissues and disease interactions. Fusarium wilt is a devastating disease of common bean caused by Fusarium oxysporum f. sp. phaseoli. In this study, we evaluated a peroxidase gene, PvPOX1, from a resistant common bean genotype, CAAS260205 and provided direct evidence for PvPOX1's role in resistance by transforming the resistant allele into a susceptible common bean genotype, BRB130, via hairy root transformation using Agrobacterium rhizogenes. Analysis of PvPOX1 gene over-expressing hairy roots showed it increased resistance to Fusarium wilt both in the roots and the rest of transgenic plants. Meanwhile, the PvPOX1 expressive level, the peroxidase activity and hydrogen peroxide (H 2 O 2 ) accumulation were also enhanced in the interaction. The result showed that the PvPOX1 gene played an essential role in Fusarium wilt resistance through the occurrence of reactive oxygen species (ROS) induced hypersensitive response. Therefore, PvPOX1 expression was proven to be a valuable gene for further analysis which can strengthen host defense response against Fusarium wilt through a ROS activated resistance mechanism. Copyright © 2017 Elsevier B.V. All rights reserved.

Triggering of the TRPV2 channel by cannabidiol sensitizes glioblastoma cells to cytotoxic chemotherapeutic agents.

PubMed

Nabissi, Massimo; Morelli, Maria Beatrice; Santoni, Matteo; Santoni, Giorgio

2013-01-01

The aggressive behavior of Glioblastoma multiforme (GBM) is mainly due to high invasiveness and proliferation rate as well as to high resistance to standard chemotherapy. Several chemotherapeutic agents like temozolomide (TMZ), carmustine (BCNU) or doxorubicin (DOXO) have been employed for treatment of GBM, but they display limited efficacy. Therefore, it is important to identify new treatment modalities to improve therapeutic effects and enhance GBM chemosensitivity. Recently, activation of the transient receptor potential vanilloid type 2 (TRPV2) has been found to inhibit human GBM cell proliferation and overcome BCNU resistance of GBM cells. Herein, we evaluated the involvement of cannabidiol (CBD)-induced TRPV2 activation, in the modulation of glioma cell chemosensitivity to TMZ, BCNU and DOXO. We found that CBD increases TRPV2 expression and activity. CBD by triggering TRPV2-dependent Ca(2+) influx increases drug uptake and synergizes with cytotoxic agents to induce apoptosis of glioma cells, whereas no effects were observed in normal human astrocytes. Moreover, as the pore region of transient receptor potential (TRP) channels is critical for ion channel permeation, we demonstrated that deletion of TRPV2 poredomain inhibits CBD-induced Ca(2+) influx, drug uptake and cytotoxic effects. Overall, we demonstrated that co-administration of cytotoxic agents together with the TRPV2 agonist CBD increases drug uptake and parallelly potentiates cytotoxic activity in human glioma cells.

Dietary polyphenols influence antimetabolite agents: methotrexate, 6-mercaptopurine and 5-fluorouracil in leukemia cell lines

PubMed Central

Mahbub, Amani; Le Maitre, Christine; Haywood-Small, Sarah; Cross, Neil; Jordan-Mahy, Nicola

2017-01-01

Polyphenols have been previously shown to sensitize leukemia cell lines to topoisomerase inhibitors. Here, we assess the effects of five polyphenols when used alone and in combination with antimetabolites: methotrexate, 6-mercaptopurine and 5-fluorouracil; in lymphoid and myeloid leukemia cells lines, and non-tumor control cells. The effects of combined treatments were investigated on ATP and glutathione levels, cell-cycle progression, DNA damage and apoptosis. Polyphenols antagonized methotrexate and 6-mercaptopurine induced cell-cycle arrest and apoptosis in most leukemia cell lines. This was associated with reduced DNA damage and increased glutathione levels, greater than that seen following individual treatments alone. In contrast, 5-fluorouracil when combined with quercetin, apigenin and rhein caused synergistic decrease in ATP levels, induction of cell-cycle arrest and apoptosis in some leukemia cell lines. However, antagonistic effects were observed when 5-fluorouracil was combined with rhein and cis-stilbene in myeloid cell lines. The effects were dependant on polyphenol type and chemotherapy agent investigated, and cell type treated. Interestingly treatment of non-tumor control cells with polyphenols protected cells from antimetabolite treatments. This suggests that polyphenols modulate the action of antimetabolite agents; more importantly they antagonized methotrexate and 6-mercaptopurine actions, thus suggesting the requirement of polyphenol-exclusion during their use. PMID:29285220

Cellular response to alkylating agent MNNG is impaired in STAT1-deficients cells.

PubMed

Ah-Koon, Laurent; Lesage, Denis; Lemadre, Elodie; Souissi, Inès; Fagard, Remi; Varin-Blank, Nadine; Fabre, Emmanuelle E; Schischmanoff, Olivier

2016-10-01

The SN 1 alkylating agents activate the mismatch repair system leading to delayed G2 /M cell cycle arrest and DNA repair with subsequent survival or cell death. STAT1, an anti-proliferative and pro-apoptotic transcription factor is known to potentiate p53 and to affect DNA-damage cellular response. We studied whether STAT1 may modulate cell fate following activation of the mismatch repair system upon exposure to the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Using STAT1-proficient or -deficient cell lines, we found that STAT1 is required for: (i) reduction in the extent of DNA lesions, (ii) rapid phosphorylation of T68-CHK2 and of S15-p53, (iii) progression through the G2 /M checkpoint and (iv) long-term survival following treatment with MNNG. Presence of STAT1 is critical for the formation of a p53-DNA complex comprising: STAT1, c-Abl and MLH1 following exposure to MNNG. Importantly, presence of STAT1 allows recruitment of c-Abl to p53-DNA complex and links c-Abl tyrosine kinase activity to MNNG-toxicity. Thus, our data highlight the important modulatory role of STAT1 in the signalling pathway activated by the mismatch repair system. This ability of STAT1 to favour resistance to MNNG indicates the targeting of STAT1 pathway as a therapeutic option for enhancing the efficacy of SN1 alkylating agent-based chemotherapy. © 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Reactive Oxygen Species (ROS) Inducible DNA Cross-Linking Agents and Their Effect on Cancer Cells and Normal Lymphocytes

PubMed Central

2015-01-01

Reducing host toxicity is one of the main challenges of cancer chemotherapy. Many tumor cells contain high levels of ROS that make them distinctively different from normal cells. We report a series of ROS-activated aromatic nitrogen mustards that selectively kill chronic lymphocytic leukemia (CLL) over normal lymphocytes. These agents showed powerful DNA cross-linking abilities when coupled with H2O2, one of the most common ROS in cancer cells, whereas little DNA cross-linking was detected without H2O2. Consistent with chemistry observation, in vitro cytotoxicity assay demonstrated that these agents induced 40–80% apoptosis in primary leukemic lymphocytes isolated from CLL patients but less than 25% cell death to normal lymphocytes from healthy donors. The IC50 for the most potent compound (2) was ∼5 μM in CLL cells, while the IC50 was not achieved in normal lymphocytes. Collectively, these data provide utility and selectivity of these agents that will inspire further and effective applications. PMID:24801734

Dependence of the cytotoxicity of DNA-damaging agents on the mismatch repair status of human cells.

PubMed

Papouli, Efterpi; Cejka, Petr; Jiricny, Josef

2004-05-15

Mismatch repair (MMR) deficiency was reported to increase resistance of mammalian cells to killing by several genotoxic substances. However, although MMR-deficient cells are approximately 100-fold more resistant to killing by S(N)1 type methylating agents than MMR-proficient controls, the sensitivity differences reported for the other agents were typically <2-fold. To test whether these differences were linked to factors other than MMR status, we studied the cytotoxicities of mitomycin C, chloroethylcyclohexyl nitrosourea, melphalan, psoralen-UVA, etoposide, camptothecin, ionizing radiation, and cis-dichlorodiaminoplatinum (cisplatin) in a strictly isogenic system. We now report that MMR deficiency reproducibly desensitized cells solely to cisplatin.

Highly Adaptable Triple-Negative Breast Cancer Cells as a Functional Model for Testing Anticancer Agents

PubMed Central

Singh, Balraj; Shamsnia, Anna; Raythatha, Milan R.; Milligan, Ryan D.; Cady, Amanda M.; Madan, Simran; Lucci, Anthony

2014-01-01

A major obstacle in developing effective therapies against solid tumors stems from an inability to adequately model the rare subpopulation of panresistant cancer cells that may often drive the disease. We describe a strategy for optimally modeling highly abnormal and highly adaptable human triple-negative breast cancer cells, and evaluating therapies for their ability to eradicate such cells. To overcome the shortcomings often associated with cell culture models, we incorporated several features in our model including a selection of highly adaptable cancer cells based on their ability to survive a metabolic challenge. We have previously shown that metabolically adaptable cancer cells efficiently metastasize to multiple organs in nude mice. Here we show that the cancer cells modeled in our system feature an embryo-like gene expression and amplification of the fat mass and obesity associated gene FTO. We also provide evidence of upregulation of ZEB1 and downregulation of GRHL2 indicating increased epithelial to mesenchymal transition in metabolically adaptable cancer cells. Our results obtained with a variety of anticancer agents support the validity of the model of realistic panresistance and suggest that it could be used for developing anticancer agents that would overcome panresistance. PMID:25279830

The antineoplastic agent α-bisabolol promotes cell death by inducing pores in mitochondria and lysosomes.

PubMed

Rigo, Antonella; Vinante, Fabrizio

2016-08-01

The sesquiterpene α-bisabolol (α-BSB) has been shown to be an effective cytotoxic agent for a variety of human cancer cells in culture and animal models. However, much of its intracellular action remains elusive. We evaluated the cytotoxic action of α-BSB against CML-T1, Jurkat and HeLa cell lines, as preclinical models for myeloid, lymphoid and epithelial neoplasias. The approach included single cell analysis (flow cytometry, immunocytology) combined with cytotoxicity and proliferation assays to characterize organelle damage, autophagy, cytostatic effect, and apoptosis. The study focuses on the relevant steps in the cytotoxic cascade triggered by α-BSB: (1) the lipid rafts through which α-BSB enters the cells, (2) the opening of pores in the mitochondria and lysosomes, (3) the activation of both caspase-dependent and caspase-independent cell death pathways, (4) the induction of autophagy and (5) apoptosis. The effectiveness of α-BSB as an agent against tumor cells is grounded on its capability to act on different layers of cell regulation to elicit different concurrent death signals, thereby neutralizing a variety of aberrant survival mechanisms leading to treatment resistance in neoplastic cell.

Highly adaptable triple-negative breast cancer cells as a functional model for testing anticancer agents.

PubMed

Singh, Balraj; Shamsnia, Anna; Raythatha, Milan R; Milligan, Ryan D; Cady, Amanda M; Madan, Simran; Lucci, Anthony

2014-01-01

A major obstacle in developing effective therapies against solid tumors stems from an inability to adequately model the rare subpopulation of panresistant cancer cells that may often drive the disease. We describe a strategy for optimally modeling highly abnormal and highly adaptable human triple-negative breast cancer cells, and evaluating therapies for their ability to eradicate such cells. To overcome the shortcomings often associated with cell culture models, we incorporated several features in our model including a selection of highly adaptable cancer cells based on their ability to survive a metabolic challenge. We have previously shown that metabolically adaptable cancer cells efficiently metastasize to multiple organs in nude mice. Here we show that the cancer cells modeled in our system feature an embryo-like gene expression and amplification of the fat mass and obesity associated gene FTO. We also provide evidence of upregulation of ZEB1 and downregulation of GRHL2 indicating increased epithelial to mesenchymal transition in metabolically adaptable cancer cells. Our results obtained with a variety of anticancer agents support the validity of the model of realistic panresistance and suggest that it could be used for developing anticancer agents that would overcome panresistance.

Effects of chemical and physical agents on recombination events in cells of the germ line of male and female Drosophila melanogaster.

PubMed

Würgler, F E

1991-01-01

Genotoxic agents can induce mutations as well as recombination in the genetic material. The fruit fly Drosophila melanogaster was one of the first assay systems to test physical and chemical agents for recombinogenic effects. Such effects can be observed in cells of the germ line as well as in somatic cells. At present information is available on 54 agents, among them 48 chemicals that have been tested in cells of the germ line of males and/or females. Effects on meiotic recombination in female germ cells cannot simply be classified as positive or negative since for a number of agents, depending on the chromosome region studied, recombination frequencies may be increased, unaffected or decreased. The male germ line of D. melanogaster represents a unique situation because meiotic recombination does not occur. Among 25 agents tested in male germ cells 24 did induce male recombination, among them alkylating, intercalating and cross-linking agents, direct-acting ones as well as compounds needing metabolic activation. With several compounds the frequency of induced recombination is highest in the heterochromatic regions near the centromeres. In brood pattern analyses, e.g., after exposure of adult males to ionizing radiation, the first appearance of crossover progeny is indicative of the sampling of exposed spermatocytes. In premeiotic cells of the male and the female germ line mitotic recombination can occur. Upon clonal expansion of the recombinant cells, clusters of identical crossovers can be observed.

Effect of Rebamipide, a Novel Antiulcer Agent, on Helicobacter pylori Adhesion to Gastric Epithelial Cells

PubMed Central

Hayashi, Shunji; Sugiyama, Toshiro; Amano, Ken-Ichi; Isogai, Hiroshi; Isogai, Emiko; Aihara, Miki; Kikuchi, Mikio; Asaka, Masahiro; Yokota, Kenji; Oguma, Keiji; Fujii, Nobuhiro; Hirai, Yoshikazu

1998-01-01

Helicobacter pylori is a major etiological agent in gastroduodenal disorders. The adhesion of H. pylori to human gastric epithelial cells is the initial step of H. pylori infection. Inhibition of H. pylori adhesion is thus a therapeutic target in the prevention of H. pylori infection. Experiments were performed to evaluate the effect of rebamipide, a novel antiulcer agent, on H. pylori adhesion to gastric epithelial cells. MKN-28 and MKN-45 cells, derived from human gastric carcinomas, were used as target cells. Ten H. pylori strains isolated from patients with chronic gastritis and gastric ulcer were used in the study. We evaluated the effect of rebamipide on H. pylori adhesion to MKN-28 and MKN-45 cells quantitatively using our previously established enzyme-linked immunosorbent assay. The adhesion of H. pylori to MKN-28 and MKN-45 cells was significantly inhibited by pretreatment of these cells with 100 μg of rebamipide per ml. However, the adhesion was not affected by the pretreatment of H. pylori with rebamipide. On the other hand, the viabilities of H. pylori, MKN-28 cells, and MKN-45 cells were not affected by rebamipide. Our studies suggest that rebamipide inhibits the adhesion of H. pylori to gastric epithelial cells. PMID:9687380

Tumor Lysing Genetically Engineered T Cells Loaded with Multi-Modal Imaging Agents

NASA Astrophysics Data System (ADS)

Bhatnagar, Parijat; Alauddin, Mian; Bankson, James A.; Kirui, Dickson; Seifi, Payam; Huls, Helen; Lee, Dean A.; Babakhani, Aydin; Ferrari, Mauro; Li, King C.; Cooper, Laurence J. N.

2014-03-01

Genetically-modified T cells expressing chimeric antigen receptors (CAR) exert anti-tumor effect by identifying tumor-associated antigen (TAA), independent of major histocompatibility complex. For maximal efficacy and safety of adoptively transferred cells, imaging their biodistribution is critical. This will determine if cells home to the tumor and assist in moderating cell dose. Here, T cells are modified to express CAR. An efficient, non-toxic process with potential for cGMP compliance is developed for loading high cell number with multi-modal (PET-MRI) contrast agents (Super Paramagnetic Iron Oxide Nanoparticles - Copper-64; SPION-64Cu). This can now be potentially used for 64Cu-based whole-body PET to detect T cell accumulation region with high-sensitivity, followed by SPION-based MRI of these regions for high-resolution anatomically correlated images of T cells. CD19-specific-CAR+SPIONpos T cells effectively target in vitro CD19+ lymphoma.

Bortezomib and Filgrastim in Promoting Stem Cell Mobilization in Patients With Non-Hodgkin Lymphoma or Multiple Myeloma Undergoing Stem Cell Transplant

ClinicalTrials.gov

2017-05-23

Adult Grade III Lymphomatoid Granulomatosis; B-cell Chronic Lymphocytic Leukemia; Contiguous Stage II Adult Burkitt Lymphoma; Contiguous Stage II Adult Diffuse Large Cell Lymphoma; Contiguous Stage II Adult Diffuse Mixed Cell Lymphoma; Contiguous Stage II Adult Diffuse Small Cleaved Cell Lymphoma; Contiguous Stage II Adult Immunoblastic Large Cell Lymphoma; Contiguous Stage II Adult Lymphoblastic Lymphoma; Contiguous Stage II Grade 1 Follicular Lymphoma; Contiguous Stage II Grade 2 Follicular Lymphoma; Contiguous Stage II Grade 3 Follicular Lymphoma; Contiguous Stage II Mantle Cell Lymphoma; Contiguous Stage II Marginal Zone Lymphoma; Contiguous Stage II Small Lymphocytic Lymphoma; Cutaneous B-cell Non-Hodgkin Lymphoma; Extranodal Marginal Zone B-cell Lymphoma of Mucosa-associated Lymphoid Tissue; Intraocular Lymphoma; Nodal Marginal Zone B-cell Lymphoma; Noncontiguous Stage II Adult Burkitt Lymphoma; Noncontiguous Stage II Adult Diffuse Large Cell Lymphoma; Noncontiguous Stage II Adult Diffuse Mixed Cell Lymphoma; Noncontiguous Stage II Adult Diffuse Small Cleaved Cell Lymphoma; Noncontiguous Stage II Adult Immunoblastic Large Cell Lymphoma; Noncontiguous Stage II Adult Lymphoblastic Lymphoma; Noncontiguous Stage II Grade 1 Follicular Lymphoma; Noncontiguous Stage II Grade 2 Follicular Lymphoma; Noncontiguous Stage II Grade 3 Follicular Lymphoma; Noncontiguous Stage II Mantle Cell Lymphoma; Noncontiguous Stage II Marginal Zone Lymphoma; Noncontiguous Stage II Small Lymphocytic Lymphoma; Progressive Hairy Cell Leukemia, Initial Treatment; Recurrent Adult Burkitt Lymphoma; Recurrent Adult Diffuse Large Cell Lymphoma; Recurrent Adult Diffuse Mixed Cell Lymphoma; Recurrent Adult Diffuse Small Cleaved Cell Lymphoma; Recurrent Adult Grade III Lymphomatoid Granulomatosis; Recurrent Adult Immunoblastic Large Cell Lymphoma; Recurrent Adult Lymphoblastic Lymphoma; Recurrent Grade 1 Follicular Lymphoma; Recurrent Grade 2 Follicular Lymphoma; Recurrent Grade 3 Follicular

Cell-Specific Production and Antimicrobial Activity of Naphthoquinones in Roots of Lithospermum erythrorhizon1

PubMed Central

Brigham, Lindy A.; Michaels, Paula J.; Flores, Hector E.

1999-01-01

Pigmented naphthoquinone derivatives of shikonin are produced at specific times and in specific cells of Lithospermum erythrorhizon roots. Normal pigment development is limited to root hairs and root border cells in hairy roots grown on “noninducing” medium, whereas induction of additional pigment production by abiotic (CuSO4) or biotic (fungal elicitor) factors increases the amount of total pigment, changes the ratios of derivatives produced, and initiates production of pigment de novo in epidermal cells. When the biological activity of these compounds was tested against soil-borne bacteria and fungi, a wide range of sensitivity was recorded. Acetyl-shikonin and β-hydroxyisovaleryl-shikonin, the two most abundant derivatives in both Agrobacterium rhizogenes-transformed “hairy-root” cultures and greenhouse-grown plant roots, were the most biologically active of the seven compounds tested. Hyphae of the pathogenic fungi Rhizoctonia solani, Pythium aphanidermatum, and Nectria hematococca induced localized pigment production upon contact with the roots. Challenge by R. solani crude elicitor increased shikonin derivative production 30-fold. We have studied the regulation of this suite of related, differentially produced, differentially active compounds to understand their role(s) in plant defense at the cellular level in the rhizosphere. PMID:9952436

Expression of cardiac function genes in adult stem cells is increased by treatment with nitric oxide agents.

PubMed

Rebelatto, Carmen K; Aguiar, Alessandra M; Senegaglia, Alexandra C; Aita, Carlos M; Hansen, Paula; Barchiki, Fabiane; Kuligovski, Crisciele; Olandoski, Márcia; Moutinho, José A; Dallagiovanna, Bruno; Goldenberg, Samuel; Brofman, Paulo S; Nakao, Lia S; Correa, Alejandro

2009-01-16

Mesenchymal stem cells (MSCs) have received special attention for cardiomyoplasty because several studies have shown that they differentiate into cardiomyocytes both in vitro and in vivo. Nitric oxide (NO) is a free radical signaling molecule that regulates several differentiation processes including cardiomyogenesis. Here, we report an investigation of the effects of two NO agents (SNAP and DEA/NO), able to activate both cGMP-dependent and -independent pathways, on the cardiomyogenic potential of bone marrow-derived mesenchymal stem cells (BM-MSCs) and adipose tissue-derived stem cells (ADSCs). The cells were isolated, cultured and treated with NO agents. Cardiac- and muscle-specific gene expression was analyzed by indirect immunofluorescence, flow cytometry, RT-PCR and real-time PCR. We found that untreated (control) ADSCs and BM-MSCs expressed some muscle markers and NO-derived intermediates induce an increased expression of some cardiac function genes in BM-MSCs and ADSCs. Moreover, NO agents considerably increased the pro-angiogenic potential mostly of BM-MSCs as determined by VEGF mRNA levels.

Immunomodulatory action of SGI-110, a hypomethylating agent, in acute myeloid leukemia cells

PubMed Central

Srivastava, Pragya; Paluch, Benjamin E.; Matsuzaki, Junko; James, Smitha R.; Collamat-Lai, Golda; Karbach, Julia; Nemeth, Michael J.; Taverna, Pietro; Karpf, Adam R.; Griffiths, Elizabeth A.

2017-01-01

The mechanism of clinical action for the FDA approved hypomethylating drugs azacitidine and decitabine remains unresolved and in this context the potential immunomodulatory effect of these agents on leukemic cells is an area of active investigation. Induced expression of methylated Cancer Testis Antigen (CTA) genes has been demonstrated in leukemic cell lines following exposure to hypomethylating drugs in vitro. SGI-110 is a novel hypomethylating dinucleotide with prolonged in vivo exposure and clinical activity in patients with MDS and AML. We demonstrate that this agent, like decitabine, produces robust re-expression of the CTAs NY-ESO-1 and MAGE-A, both in vitro and in leukemia-bearing AML xenografts. Upregulation of these genes in vitro was sufficient to induce cytotoxicity by HLA-compatible CD8+ T-cells specific for NY-ESO-1, a well-recognized and immunogenic CTA. Additionally, exposure to SGI-110 enhances MHC class I and co-stimulatory molecule expression, potentially contributing to recognition of CTAs. SGI-110, like the parent compound decitabine, induces expression of CTAs and might modulate immune recognition of myeloid malignancy. PMID:25260825

Alkylating chemotherapeutic agents cyclophosphamide and melphalan cause functional injury to human bone marrow-derived mesenchymal stem cells.

PubMed

Kemp, Kevin; Morse, Ruth; Sanders, Kelly; Hows, Jill; Donaldson, Craig

2011-07-01

The adverse effects of melphalan and cyclophosphamide on hematopoietic stem cells are well-known; however, the effects on the mesenchymal stem cells (MSCs) residing in the bone marrow are less well characterised. Examining the effects of chemotherapeutic agents on patient MSCs in vivo is difficult due to variability in patients and differences in the drug combinations used, both of which could have implications on MSC function. As drugs are not commonly used as single agents during high-dose chemotherapy (HDC) regimens, there is a lack of data comparing the short- or long-term effects these drugs have on patients post treatment. To help address these problems, the effects of the alkylating chemotherapeutic agents cyclophosphamide and melphalan on human bone marrow MSCs were evaluated in vitro. Within this study, the exposure of MSCs to the chemotherapeutic agents cyclophosphamide or melphalan had strong negative effects on MSC expansion and CD44 expression. In addition, changes were seen in the ability of MSCs to support hematopoietic cell migration and repopulation. These observations therefore highlight potential disadvantages in the use of autologous MSCs in chemotherapeutically pre-treated patients for future therapeutic strategies. Furthermore, this study suggests that if the damage caused by chemotherapeutic agents to marrow MSCs is substantial, it would be logical to use cultured allogeneic MSCs therapeutically to assist or repair the marrow microenvironment after HDC.

Risk Mitigation in Preventing Adventitious Agent Contamination of Mammalian Cell Cultures.

PubMed

Shiratori, Masaru; Kiss, Robert

2017-11-14

Industrial-scale mammalian cell culture processes have been contaminated by viruses during the culturing phase. Although the historical frequency of such events has been quite low, the impact of contamination can be significant for the manufacturing company and for the supply of the product to patients. This chapter discusses sources of adventitious agent contamination risk in a cell culture process, provides a semiquantitative assessment of such risks, and describes potential process barriers that can be used to reduce contamination risk. High-temperature, short-time (HTST) heat treatment is recommended as the process barrier of choice, when compatible with the process. A case study assessing the compatibility of HTST heat treatment with a cell culture medium is presented, and lessons learned are shared from our experiences over many years of developing and implementing virus barriers in mammalian cell culture processes. Graphical Abstract.

[Relationship between sensitivity of tumor cells to chemotherapeutic agent in vivo and in vitro: experiment with mouse lymphoma cells].

PubMed

Li, Chuan-gang; Li, Mo-lin; Shu, Xiao-hong; Jia, Yu-jie; Liu, Yong-ji; Li, Ming

2007-06-12

To study the relationship of the sensitivity of tumor cells to chemotherapeutic agent between in vivo and in vitro. Mouse lymphoma cells of the line E14 were cultured and melphalan resistant EL4 cell line (EL4/melphalan) was established by culturing EL4 cells with continuous low-concentration and intermittent gradually-increasing-concentration of melphalan in vitro. MTT assay was used to evaluate the drug sensitivity and the resistance index of the EL4/melphalan cells to melphalan was calculated. EL4/melphalan and EL4 cells of the concentration of 5 x 10(8)/L were inoculated separately into 20 C57BL/6 mice subcutaneously. 12 days later, the EL4 and EL4/melphalan tumor-bearing mice were randomly divided into 2 groups respectively, 5 mice in each group. Treatment groups were given 7.5 mg/kg melphalan intraperitoneally, and control groups were given the same volume of normal saline. The tumor size was observed every other day. Compared with the EL4 cells, the EL4/melphalan cells had no obvious changes morphologically. They could grow in RPMI 1640 medium containing 5 mg/ml melphalan. The resistance index was 2.87 against melphalan. After the treatment of melphalan of the dose 7.5 mg/kg, the tumor sizes of the treatment groups and control groups inoculated with both EL4 cells and the EL4/melphalan cells gradually decreased at the similar speed, and about one week later all tumors disappeared. However, the tumors of the control groups grew progressively and all the mice died at last. The chemotherapeutic effects of tumors in vivo have nothing to do with the effects of the chemotherapeutic agents on tumor cells in vitro. The tumor cells resistant to melphalan in vitro remain sensitive to the drug in vivo.

A unique 19F MRI agent for the tracking of non phagocytic cells in vivo.

PubMed

Moonshi, Shehzahdi S; Zhang, Cheng; Peng, Hui; Puttick, Simon; Rose, Stephen; Fisk, Nicholas M; Bhakoo, Kishore; Stringer, Brett W; Qiao, Greg G; Gurr, Paul A; Whittaker, Andrew K

2018-05-03

There is currently intense interest in new methods for understanding the fate of therapeutically-relevant cells, such as mesenchymal stem cells (MSCs). The absence of a confounding background signal and consequent unequivocal assignment makes 19F MRI one of the most attractive modalities for the tracking of injected cells in vivo. We describe here the synthesis of novel partly-fluorinated polymeric nanoparticles with small size and high fluorine content as MRI agents. The polymers, constructed from perfluoropolyether methacrylate (PFPEMA) and oligo(ethylene glycol) methacrylate (OEGMA) have favourable cell uptake profiles and excellent MRI performance. To facilitate cell studies the polymer was further conjugated with a fluorescent dye creating a dual-modal imaging agent. The efficacy of labelling of MSCs was assessed using 19F NMR, flow cytometry and confocal microscopy. The labelling efficiency of 2.6 ± 0.1 × 1012 19F atoms per cell, and viability of >90% demonstrates high uptake and good tolerance by the cells. This loading translates to a minimum 19F MRI detection sensitivity of ∼7.4 × 103 cells per voxel. Importantly, preliminary in vivo data demonstrate that labelled cells can be readily detected within a short acquisition scan period (12 minutes). Hence, these copolymers show outstanding potential for 19F MRI cellular tracking and for quantification of non-phagocytic and therapeutically-relevant cells in vivo.

Understanding to Hierarchical Microstructures of Crab (Chinese hairy) Shell as a Natural Architecture

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chuanqiang, Zhou; Xiangxiang, Gong; School of Chemistry and Chemical Engineering, Yangzhou University, Yangzhou

This work was done to better understand the microstructures, composition and mechanical properties of Chinese hairy crab shell. For fully revealing its hierarchical microstructure, the crab shell was observed with electron microscope under different magnifications from different facets. XRD, EDS, FTIR and TGA techniques have been used to characterize the untreated and chemically-treated crab shells, which provided enough information to determine the species and relative content of components in this biomaterial. Combined the microstructures with constituents analysis, the structural principles of crab shell was detailedly realized from different structural levels beyond former reports. To explore the relationship between structure andmore » function, the mechanical properties of shell have been measured through performing tensile tests. The contributions of organics and minerals in shell to the mechanical properties were also discussed by measuring the tensile strength of de-calcification samples treated with HCl solution.« less

Racial Differences in the Overall Survival of Hairy Cell Leukemia in the United States: A Population-Based Analysis of the Surveillance, Epidemiology, and End Results Database.

PubMed

Giri, Smith; Shrestha, Rajesh; Pathak, Ranjan; Bhatt, Vijaya Raj

2015-08-01

Several studies have reported excellent long-term overall survival (OS) of patients with hairy cell leukemia (HCL) without racial disparity. Studies in other cancers have demonstrated worse mortality among African American (AA) individuals. We used the Surveillance, Epidemiology, and End Results 18 database to identify HCL patients diagnosed between 1978 and 2011. Kaplan-Meier curves were plotted to estimate OS. Univariate analysis using the life table method and multivariate Cox regression model were used to determine the independent effect of race on OS. The study population included 78% men and had a median age of 56 years. Race included 93% white, 3.5% Asian/Pacific Islander, and 3.5% AA. The 10-year OS was significantly less for AA as compared with white and Asian/Pacific Islander individuals (54% vs. 72% vs. 75%; P < .001). A Kaplan-Meier survival curve showed a significantly worse OS for AA versus other races (P < .001). In a multivariate analysis, AA race remained an independent predictor for a worse OS (hazard ratio 1.77; 95% confidence interval, 1.30-2.40; P < .001) after adjusting for age, sex, year of diagnosis, and marital status. In this population-based study, only half of AA patients but more than two-thirds of HCL patients from other racial groups were alive at 10 years. Such drastic racial differences in OS of HCL patients at the population level mandates further evaluation of the contributory biological, socioeconomic, health system, and other factors. Understanding and overcoming such racial disparities might close the racial differences in OS of this potentially curable disease. Copyright © 2015 Elsevier Inc. All rights reserved.

The c4h, tat, hppr and hppd Genes Prompted Engineering of Rosmarinic Acid Biosynthetic Pathway in Salvia miltiorrhiza Hairy Root Cultures

PubMed Central

Gao, Shouhong; Saechao, Saengking; Di, Peng; Chen, Junfeng; Chen, Wansheng

2011-01-01

Rational engineering to produce biologically active plant compounds has been greatly impeded by our poor understanding of the regulatory and metabolic pathways underlying the biosynthesis of these compounds. Here we capitalized on our previously described gene-to-metabolite network in order to engineer rosmarinic acid (RA) biosynthesis pathway for the production of beneficial RA and lithospermic acid B (LAB) in Salvia miltiorrhiza hairy root cultures. Results showed their production was greatly elevated by (1) overexpression of single gene, including cinnamic acid 4-hydroxylase (c4h), tyrosine aminotransferase (tat), and 4-hydroxyphenylpyruvate reductase (hppr), (2) overexpression of both tat and hppr, and (3) suppression of 4-hydroxyphenylpyruvate dioxygenase (hppd). Co-expression of tat/hppr produced the most abundant RA (906 mg/liter) and LAB (992 mg/liter), which were 4.3 and 3.2-fold more than in their wild-type (wt) counterparts respectively. And the value of RA concentration was also higher than that reported before, that produced by means of nutrient medium optimization or elicitor treatment. It is the first report of boosting RA and LAB biosynthesis through genetic manipulation, providing an effective approach for their large-scale commercial production by using hairy root culture systems as bioreactors. PMID:22242141

Effect of sulphur mustard on human skin cell lines with differential agent sensitivity.

PubMed

Simpson, Rachel; Lindsay, Christopher D

2005-01-01

The ability of sulphur mustard (HD) to induce DNA damage places limits on the efficacy of approaches aimed at protecting human cells from the cytotoxic effects of HD using a variety of protective agents such as thiol-containing esters and protease inhibitors. In the present study, potential alternative strategies were investigated by examining the differential effects of HD on G361, SVK14, HaCaT and NCTC 2544 human skin cells. The G361 cell line was more resistant to the cytotoxic effects of HD than the NCTC, HaCaT and SVK14 cell lines at HD doses of >3 and <100 microM HD as determined by the MTT assay. At 72 h after exposure to 60 microM HD there was up to an 8.8-fold difference (P < 0.0001) between G361 and SVK14 cell culture viability. Buthionine sulphoximine (BSO) pretreatment increased the sensitivity of all four cell lines to HD. A substantial proportion of the resistance of G361 cells to HD was attributable to BSO-mediated effects on antioxidant-mediated metabolism, although G361 cultures still retained a high degree of viability at 30 microM HD following BSO pretreatment. Cell cycle analysis confirmed that SVK14 cells were relatively more sensitive to HD, as shown by the 2.1-fold reduction (P < 0.0001) in the percentage of cells in G0/G1 phase 24 h after HD exposure compared with control cultures. This compared well with a 1.2-fold increase (P < 0.05) in the percentage of G361 cells in G0/G1 phase following HD exposure, suggesting the existence of a more efficient G0/G1 checkpoint control mechanism in this cell line. Manipulation of the cell cycle using various modulating agents did not increase the resistance of cell lines to the cytotoxic effects of HD. Crown copyright 2005

What is the role of biosimilar G-CSF agents in hematopoietic stem cell mobilization at present?

PubMed

Korkmaz, Serdal; Altuntas, Fevzi

2017-12-01

Mobilization of hematopoietic stem cells, which has largely replaced bone marrow harvesting as a source of hematopoietic stem cells, using recombinant agents such as filgrastim or lenograstim has become a standard procedure in both patients and healthy donors prior to peripheral blood stem cell collection for autologous and allogeneic stem cell transplantation. Published literature data suggest that mobilization with recombinant granulocyte-colony stimulating factor (G-CSF) is safe and mobilization outcomes are satisfactory. In recent years, besides G-CSF originators, biosimilar G-CSF agents have been approved by the regulatory agencies for the same indications. Current data showed that by using the biosimilar G-CSF, similar results regarding safety and efficacy of hematopoietic stem cell mobilization may be achieved compared to the originator G-CSF. Although the issues such as the similarity to a licenced biological medicine, differences in manufacturing processes, the potential to cause immunogenicity, extrapolation and interchangeability of these biosimilar products are still being discussed by the scientific area, however, more experience with these agents now exists in approved endications and there seems to be no reason to expect significant differences between biosimilar G-CSF and originator G-CSF regarding their efficacy and safety in both patients and healthy donors. Also, the significant cost savings of biosimilars in real life setting may enhance the use of these agents in the future. Nonetheless, the collection of long-term follow-up data is mandatory for both patients and healthy donors, and multicentre randomized clinical trials that directly compare biosimilar G-CSF with the originator G-CSF are needed in order to allow the transplant community to make informed decisions regarding the choice of G-CSF. Copyright © 2017 Elsevier Ltd. All rights reserved.

HeLa Cells Containing a Truncated Form of DNA Polymerase Beta are More Sensitized to Alkylating Agents than to Agents Inducing Oxidative Stress.

PubMed

Khanra, Kalyani; Chakraborty, Anindita; Bhattacharyya, Nandan

2015-01-01

The present study was aimed at determining the effects of alkylating and oxidative stress inducing agents on a newly identified variant of DNA polymerase beta (polβ Δ208-304) specific for ovarian cancer. Pol β Δ208-304 has a deletion of exons 11-13 which lie in the catalytic part of enzyme. We compared the effect of these chemicals on HeLa cells and HeLa cells stably transfected with this variant cloned into in pcDNAI/neo vector by MTT, colony forming and apoptosis assays. Polβ Δ208-304 cells exhibited greater sensitivity to an alkylating agent and less sensitivity towards H2O2 and UV when compared with HeLa cells alone. It has been shown that cell death in Pol β Δ208-304 transfected HeLa cells is mediated by the caspase 9 cascade. Exon 11 has nucleotidyl selection activity, while exons 12 and 13 have dNTP selection activity. Hence deletion of this part may affect polymerizing activity although single strand binding and double strand binding activity may remain same. The lack of this part may adversely affect catalytic activity of DNA polymerase beta so that the variant may act as a dominant negative mutant. This would represent clinical significance if translated into a clinical setting because resistance to radiation or chemotherapy during the relapse of the disease could be potentially overcome by this approach.

Targeting ferritin receptors for the selective delivery of imaging and therapeutic agents to breast cancer cells

NASA Astrophysics Data System (ADS)

Geninatti Crich, S.; Cadenazzi, M.; Lanzardo, S.; Conti, L.; Ruiu, R.; Alberti, D.; Cavallo, F.; Cutrin, J. C.; Aime, S.

2015-04-01

In this work the selective uptake of native horse spleen ferritin and apoferritin loaded with MRI contrast agents has been assessed in human breast cancer cells (MCF-7 and MDA-MB-231). The higher expression of L-ferritin receptors (SCARA5) led to an enhanced uptake in MCF-7 as shown in T2 and T1 weighted MR images, respectively. The high efficiency of ferritin internalization in MCF-7 has been exploited for the simultaneous delivery of curcumin, a natural therapeutic molecule endowed with antineoplastic and anti-inflammatory action, and the MRI contrast agent Gd-HPDO3A. This theranostic system is able to treat selectively breast cancer cells over-expressing ferritin receptors. By entrapping in apoferritin both Gd-HPDO3A and curcumin, it was possible to deliver a therapeutic dose of 167 μg ml-1 (as calculated by MRI) of this natural drug to MCF-7 cells, thus obtaining a significant reduction of cell proliferation.In this work the selective uptake of native horse spleen ferritin and apoferritin loaded with MRI contrast agents has been assessed in human breast cancer cells (MCF-7 and MDA-MB-231). The higher expression of L-ferritin receptors (SCARA5) led to an enhanced uptake in MCF-7 as shown in T2 and T1 weighted MR images, respectively. The high efficiency of ferritin internalization in MCF-7 has been exploited for the simultaneous delivery of curcumin, a natural therapeutic molecule endowed with antineoplastic and anti-inflammatory action, and the MRI contrast agent Gd-HPDO3A. This theranostic system is able to treat selectively breast cancer cells over-expressing ferritin receptors. By entrapping in apoferritin both Gd-HPDO3A and curcumin, it was possible to deliver a therapeutic dose of 167 μg ml-1 (as calculated by MRI) of this natural drug to MCF-7 cells, thus obtaining a significant reduction of cell proliferation. Electronic supplementary information (ESI) available: Competition studies with free apoferritin, Fig. S1; APO-FITC intracellular distribution by

Lysosomotropic properties of weakly basic anticancer agents promote cancer cell selectivity in vitro.

PubMed

Ndolo, Rosemary A; Luan, Yepeng; Duan, Shaofeng; Forrest, M Laird; Krise, Jeffrey P

2012-01-01

Drug distribution in cells is a fundamentally important, yet often overlooked, variable in drug efficacy. Many weakly basic anticancer agents accumulate extensively in the acidic lysosomes of normal cells through ion trapping. Lysosomal trapping reduces the activity of anticancer drugs, since anticancer drug targets are often localized in the cell cytosol or nucleus. Some cancer cells have defective acidification of lysosomes, which causes a redistribution of trapped drugs from the lysosomes to the cytosol. We have previously established that such differences in drug localization between normal and cancer cells can contribute to the apparent selectivity of weakly basic drugs to cancer cells in vitro. In this work, we tested whether this intracellular distribution-based drug selectivity could be optimized based on the acid dissociation constant (pKa) of the drug, which is one of the determinants of lysosomal sequestration capacity. We synthesized seven weakly basic structural analogs of the Hsp90 inhibitor geldanamycin (GDA) with pKa values ranging from 5 to 12. The selectivity of each analog was expressed by taking ratios of anti-proliferative IC(50) values of the inhibitors in normal fibroblasts to the IC(50) values in human leukemic HL-60 cells. Similar selectivity assessments were performed in a pair of cancer cell lines that differed in lysosomal pH as a result of siRNA-mediated alteration of vacuolar proton ATPase subunit expression. Optimal selectivity was observed for analogs with pKa values near 8. Similar trends were observed with commercial anticancer agents with varying weakly basic pKa values. These evaluations advance our understanding of how weakly basic properties can be optimized to achieve maximum anticancer drug selectivity towards cancer cells with defective lysosomal acidification in vitro. Additional in vivo studies are needed to examine the utility of this approach for enhancing selectivity.

Lysosomotropic Properties of Weakly Basic Anticancer Agents Promote Cancer Cell Selectivity In Vitro

PubMed Central

Ndolo, Rosemary A.; Luan, Yepeng; Duan, Shaofeng; Forrest, M. Laird; Krise, Jeffrey P.

2012-01-01

Drug distribution in cells is a fundamentally important, yet often overlooked, variable in drug efficacy. Many weakly basic anticancer agents accumulate extensively in the acidic lysosomes of normal cells through ion trapping. Lysosomal trapping reduces the activity of anticancer drugs, since anticancer drug targets are often localized in the cell cytosol or nucleus. Some cancer cells have defective acidification of lysosomes, which causes a redistribution of trapped drugs from the lysosomes to the cytosol. We have previously established that such differences in drug localization between normal and cancer cells can contribute to the apparent selectivity of weakly basic drugs to cancer cells in vitro. In this work, we tested whether this intracellular distribution-based drug selectivity could be optimized based on the acid dissociation constant (pKa) of the drug, which is one of the determinants of lysosomal sequestration capacity. We synthesized seven weakly basic structural analogs of the Hsp90 inhibitor geldanamycin (GDA) with pKa values ranging from 5 to 12. The selectivity of each analog was expressed by taking ratios of anti-proliferative IC50 values of the inhibitors in normal fibroblasts to the IC50 values in human leukemic HL-60 cells. Similar selectivity assessments were performed in a pair of cancer cell lines that differed in lysosomal pH as a result of siRNA-mediated alteration of vacuolar proton ATPase subunit expression. Optimal selectivity was observed for analogs with pKa values near 8. Similar trends were observed with commercial anticancer agents with varying weakly basic pKa values. These evaluations advance our understanding of how weakly basic properties can be optimized to achieve maximum anticancer drug selectivity towards cancer cells with defective lysosomal acidification in vitro. Additional in vivo studies are needed to examine the utility of this approach for enhancing selectivity. PMID:23145164

Cell targeting peptides as smart ligands for targeting of therapeutic or diagnostic agents: a systematic review.

PubMed

Mousavizadeh, Ali; Jabbari, Ali; Akrami, Mohammad; Bardania, Hassan

2017-10-01

Cell targeting peptides (CTP) are small peptides which have high affinity and specificity to a cell or tissue targets. They are typically identified by using phage display and chemical synthetic peptide library methods. CTPs have attracted considerable attention as a new class of ligands to delivery specifically therapeutic and diagnostic agents, because of the fact they have several advantages including easy synthesis, smaller physical sizes, lower immunogenicity and cytotoxicity and their simple and better conjugation to nano-carriers and therapeutic or diagnostic agents compared to conventional antibodies. In this systematic review, we will focus on the basic concepts concerning the use of cell-targeting peptides (CTPs), following the approaches of selecting them from peptide libraries. We discuss several developed strategies for cell-specific delivery of different cargos by CTPs, which are designed for drug delivery and diagnostic applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Cell Culture System for Analysis of Genetic Heterogeneity Within Hepatocellular Carcinomas and Response to Pharmacologic Agents.

PubMed

Gao, Qiang; Wang, Zhi-Chao; Duan, Meng; Lin, Yi-Hui; Zhou, Xue-Ya; Worthley, Daniel L; Wang, Xiao-Ying; Niu, Gang; Xia, Yuchao; Deng, Minghua; Liu, Long-Zi; Shi, Jie-Yi; Yang, Liu-Xiao; Zhang, Shu; Ding, Zhen-Bin; Zhou, Jian; Liang, Chun-Min; Cao, Ya; Xiong, Lei; Xi, Ruibin; Shi, Yong-Yong; Fan, Jia

2017-01-01

No targeted therapies have been found to be effective against hepatocellular carcinoma (HCC), possibly due to the large degree of intratumor heterogeneity. We performed genetic analyses of different regions of HCCs to evaluate levels of intratumor heterogeneity and associate alterations with responses to different pharmacologic agents. We obtained samples of HCCs (associated with hepatitis B virus infection) from 10 patients undergoing curative resection, before adjuvant therapy, at hospitals in China. We collected 4-9 spatially distinct samples from each tumor (55 regions total), performed histologic analyses, isolated cancer cells, and carried them low-passage culture. We performed whole-exome sequencing, copy-number analysis, and high-throughput screening of the cultured primary cancer cells. We tested responses of an additional 105 liver cancer cell lines to a fibroblast growth factor receptor (FGFR) 4 inhibitor. We identified a total of 3670 non-silent mutations (3192 missense, 94 splice-site variants, and 222 insertions or deletions) in the tumor samples. We observed considerable intratumor heterogeneity and branched evolution in all 10 tumors; the mean percentage of heterogeneous mutations in each tumor was 39.7% (range, 12.9%-68.5%). We found significant mutation shifts toward C>T and C>G substitutions in branches of phylogenetic trees among samples from each tumor (P < .0001). Of note, 14 of the 26 oncogenic alterations (53.8%) varied among subclones that mapped to different branches. Genetic alterations that can be targeted by existing pharmacologic agents (such as those in FGF19, DDR2, PDGFRA, and TOP1) were identified in intratumor subregions from 4 HCCs and were associated with sensitivity to these agents. However, cells from the remaining subregions, which did not have these alterations, were not sensitive to these drugs. High-throughput screening identified pharmacologic agents to which these cells were sensitive, however. Overexpression of FGF19

Enhanced Eradication of Lymphoma by Tumor-Specific Cytotoxic T Cells Secreting and Engineered Tumor-Specific Immunotoxin

DTIC Science & Technology

2009-06-01

target delivery of an immunotoxin, the CD22 -Pseudomonas exotoxin A ( CD22 -PEA), which has already been used in a clinical setting . The toxin portion...contains the transloc ating and ADP-ribosylat ing dom ains of PEA, and the native cell-binding portion is replaced with a CD22 scFv that directs...targeting to B lymphocytes. CD22 -PEA was tested in a Phase I trial in B-cell malignancies, but tumor responses, particularly in hairy cell leukemia

Spatial Control of Cell Transfection Using Soluble or Solid-Phase Redox Agents and a Redox-Active Ferrocenyl Lipid

PubMed Central

Aytar, Burcu S.; Muller, John P. E.; Kondo, Yukishige; Abbott, Nicholas L.; Lynn, David M.

2013-01-01

We report principles for active, user-defined control over the locations and timing with which DNA is expressed in cells. Our approach exploits unique properties of a ferrocenyl cationic lipid that is inactive when oxidized, but active when chemically reduced. We show that methods that exert spatial control over the administration of reducing agents can lead to local activation of lipoplexes and spatial control over gene expression. The versatility of this approach is demonstrated using both soluble and solid-phase reducing agents. These methods provide control over cell transfection, including methods for remote activation and the patterning of expression using solid-phase redox agents, that are difficult to achieve using conventional lipoplexes. PMID:23965341

Spatial control of cell transfection using soluble or solid-phase redox agents and a redox-active ferrocenyl lipid.

PubMed

Aytar, Burcu S; Muller, John P E; Kondo, Yukishige; Abbott, Nicholas L; Lynn, David M

2013-09-11

We report principles for active, user-defined control over the locations and timing with which DNA is expressed in cells. Our approach exploits unique properties of a ferrocenyl cationic lipid that is inactive when oxidized, but active when chemically reduced. We show that methods that exert spatial control over the administration of reducing agents can lead to local activation of lipoplexes and spatial control over gene expression. The versatility of this approach is demonstrated using both soluble and solid-phase reducing agents. These methods provide control over cell transfection, including methods for remote activation and the patterning of expression using solid-phase redox agents, that are difficult to achieve using conventional lipoplexes.

Cellular Immunotherapy Following Chemotherapy in Treating Patients With Recurrent Non-Hodgkin Lymphomas, Chronic Lymphocytic Leukemia or B-Cell Prolymphocytic Leukemia

ClinicalTrials.gov

2018-04-20

Post-transplant Lymphoproliferative Disorder; B-Cell Prolymphocytic Leukemia; Recurrent Adult Burkitt Lymphoma; Recurrent Adult Diffuse Large Cell Lymphoma; Recurrent Grade 1 Follicular Lymphoma; Recurrent Grade 2 Follicular Lymphoma; Recurrent Grade 3 Follicular Lymphoma; Recurrent Mantle Cell Lymphoma; Recurrent Marginal Zone Lymphoma; Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; Refractory Hairy Cell Leukemia; B-Cell Lymphoma, Unclassifiable, With Features Intermediate Between Diffuse Large B-Cell Lymphoma and Burkitt Lymphoma; B-Cell Lymphoma, Unclassifiable, With Features Intermediate Between Diffuse Large B-Cell Lymphoma and Classical Hodgkin Lymphoma; Recurrent Lymphoplasmacytic Lymphoma

Antitumor agent 25-epi Ritterostatin GN1N induces endoplasmic reticulum stress and autophagy mediated cell death in melanoma cells.

PubMed

Riaz Ahmed, Kausar Begam; Kanduluru, Ananda Kumar; Feng, Li; Fuchs, Philip L; Huang, Peng

2017-05-01

Metastatic melanoma is the most aggressive of all skin cancers and is associated with poor prognosis owing to lack of effective treatments. 25-epi Ritterostatin GN1N is a novel antitumor agent with yet undefined mechanisms of action. We sought to delineate the antitumor mechanisms of 25-epi Ritterostatin GN1N in melanoma cells to determine the potential of this compound as a treatment for melanoma. Activation of the endoplasmic reticulum (ER) stress protein glucose-regulated protein 78 (GRP78) has been associated with increased melanoma progression, oncogenic signaling, drug resistance, and suppression of cell death. We found that 25-epi Ritterostatin GN1N induced cell death in melanoma cells at nanomolar concentrations, and this cell death was characterized by inhibition of GRP78 expression, increased expression of the ER stress marker CHOP, loss of mitochondrial membrane potential, and lipidation of the autophagy marker protein LC3B. Importantly, normal melanocytes exhibited limited sensitivity to 25-epi Ritterostatin GN1N. Subsequent in vivo results demonstrated that 25-epi Ritterostatin GN1N reduced melanoma growth in mouse tumor xenografts and did not affect body weight, suggesting minimal toxicity. In summary, our findings indicate that 25-epi Ritterostatin GN1N causes ER stress and massive autophagy, leading to collapse of mitochondrial membrane potential and cell death in melanoma cells, with minimal effects in normal melanocytes. Thus, 25-epi Ritterostatin GN1N is a promising anticancer agent that warrants further investigation.

Salidroside as a Novel Protective Agent to Improve Red Blood Cell Cryopreservation.

PubMed

Alotaibi, Noha A S; Slater, Nigel K H; Rahmoune, Hassan

2016-01-01

Glycerol and trehalose have been widely examined as protective agents in the cryopreservation of red blood cells (RBCs). However, the effectiveness of these reagents alone on cell viability is moderate. Here, the addition of salidroside attenuated oxidative damage of sheep RBCs prior to and post cryostorage. The supplementation of salidroside to the cryopreservation media containing 10% glycerol improved RBC survival by approximately 61.1±4.8% vs 37.9±4.6%. A smaller effect was seen in RBCs cryopreserved in 300 mM trehalose where the addition of salidroside improved survival by 7.6±0.3%. Furthermore, the addition of salidroside to cold storage solution demonstrated a significant reduction of haemolysis after 4 days for RBCs loaded with either glycerol or trehalose, compared to cells incubated without salidroside. RBCs survival was 2-fold greater following freezing in trehalose, compared with glycerol. After 10 days, salidroside enabled a lower haemolysis of 16.7±1.3% compared to 29.0±8.4% for cells incubated without salidroside. However, salidroside had no effect on RBCs which had been frozen in glycerol as the resulting haemolysis rate by day 10 was approximately 60%. Salidroside increased glutathione reductase activity and decreased lactate dehydrogenase activity. Furthermore, it led to reduced carbonylation of proteins in both glycerol and trehalose loaded cells. Finally, no effect on lipid peroxidation was found in the glycerol loaded RBCs although this was reduced in RBCs loaded with trehalose and salidroside. The present findings confirm the potential use of salidroside as a novel protective agent in cryopreservation and refrigerated storage of sheep RBCs.

Self-assembled dual-modality contrast agents for non-invasive stem cell tracking via near-infrared fluorescence and magnetic resonance imaging.

PubMed

Liu, Hong; Tan, Yan; Xie, Lisi; Yang, Lei; Zhao, Jing; Bai, Jingxuan; Huang, Ping; Zhan, Wugen; Wan, Qian; Zou, Chao; Han, Yali; Wang, Zhiyong

2016-09-15

Stem cells hold great promise for treating various diseases. However, one of the main drawbacks of stem cell therapy is the lack of non-invasive image-tracking technologies. Although magnetic resonance imaging (MRI) and near-infrared fluorescence (NIRF) imaging have been employed to analyse cellular and subcellular events via the assistance of contrast agents, the sensitivity and temporal resolution of MRI and the spatial resolution of NIRF are still shortcomings. In this study, superparamagnetic iron oxide nanocrystals and IR-780 dyes were co-encapsulated in stearic acid-modified polyethylenimine to form a dual-modality contrast agent with nano-size and positive charge. These resulting agents efficiently labelled stem cells and did not influence the cellular viability and differentiation. Moreover, the labelled cells showed the advantages of dual-modality imaging in vivo. Copyright © 2016 Elsevier Inc. All rights reserved.

Micropropagation of Salvia wagneriana Polak and hairy root cultures with rosmarinic acid production.

PubMed

Ruffoni, Barbara; Bertoli, Alessandra; Pistelli, Laura; Pistelli, Luisa

2016-01-04

Salvia wagneriana Polak is a tropical species native to Central America, well adapted to grow in the Mediterranean basin for garden decoration. Micropropagation has been assessed from axillary shoots of adult plants using a Murashige and Skoog basal medium, with the addition of 1.33-μM 6-benzylaminopurine for shoot proliferation; the subsequent rooting phase occurred in plant growth regulator-free medium. The plants were successfully acclimatised with high survival frequency. Hairy roots were induced after co-cultivation of leaf lamina and petiole fragments with Agrobacterium rhizogenes and confirmed by PCR. The establishment and proliferation of the selected HRD3 line were obtained in hormone-free liquid medium and the production of rosmarinic acid (RA) was evaluated after elicitation. The analysis of RA was performed by LC-ESI-DAD-MS in the hydroalcoholic extracts. The addition of casein hydrolysate increased the RA production, whereas no enrichment was observed after the elicitation with jasmonic acid.

A highly reversible room-temperature lithium metal battery based on crosslinked hairy nanoparticles

NASA Astrophysics Data System (ADS)

Choudhury, Snehashis; Mangal, Rahul; Agrawal, Akanksha; Archer, Lynden A.

2015-12-01

Rough electrodeposition, uncontrolled parasitic side-reactions with electrolytes and dendrite-induced short-circuits have hindered development of advanced energy storage technologies based on metallic lithium, sodium and aluminium electrodes. Solid polymer electrolytes and nanoparticle-polymer composites have shown promise as candidates to suppress lithium dendrite growth, but the challenge of simultaneously maintaining high mechanical strength and high ionic conductivity at room temperature has so far been unmet in these materials. Here we report a facile and scalable method of fabricating tough, freestanding membranes that combine the best attributes of solid polymers, nanocomposites and gel-polymer electrolytes. Hairy nanoparticles are employed as multifunctional nodes for polymer crosslinking, which produces mechanically robust membranes that are exceptionally effective in inhibiting dendrite growth in a lithium metal battery. The membranes are also reported to enable stable cycling of lithium batteries paired with conventional intercalating cathodes. Our findings appear to provide an important step towards room-temperature dendrite-free batteries.

A highly reversible room-temperature lithium metal battery based on crosslinked hairy nanoparticles.

PubMed

Choudhury, Snehashis; Mangal, Rahul; Agrawal, Akanksha; Archer, Lynden A

2015-12-04

Rough electrodeposition, uncontrolled parasitic side-reactions with electrolytes and dendrite-induced short-circuits have hindered development of advanced energy storage technologies based on metallic lithium, sodium and aluminium electrodes. Solid polymer electrolytes and nanoparticle-polymer composites have shown promise as candidates to suppress lithium dendrite growth, but the challenge of simultaneously maintaining high mechanical strength and high ionic conductivity at room temperature has so far been unmet in these materials. Here we report a facile and scalable method of fabricating tough, freestanding membranes that combine the best attributes of solid polymers, nanocomposites and gel-polymer electrolytes. Hairy nanoparticles are employed as multifunctional nodes for polymer crosslinking, which produces mechanically robust membranes that are exceptionally effective in inhibiting dendrite growth in a lithium metal battery. The membranes are also reported to enable stable cycling of lithium batteries paired with conventional intercalating cathodes. Our findings appear to provide an important step towards room-temperature dendrite-free batteries.

A highly reversible room-temperature lithium metal battery based on crosslinked hairy nanoparticles

PubMed Central

Choudhury, Snehashis; Mangal, Rahul; Agrawal, Akanksha; Archer, Lynden A.

2015-01-01

Rough electrodeposition, uncontrolled parasitic side-reactions with electrolytes and dendrite-induced short-circuits have hindered development of advanced energy storage technologies based on metallic lithium, sodium and aluminium electrodes. Solid polymer electrolytes and nanoparticle-polymer composites have shown promise as candidates to suppress lithium dendrite growth, but the challenge of simultaneously maintaining high mechanical strength and high ionic conductivity at room temperature has so far been unmet in these materials. Here we report a facile and scalable method of fabricating tough, freestanding membranes that combine the best attributes of solid polymers, nanocomposites and gel-polymer electrolytes. Hairy nanoparticles are employed as multifunctional nodes for polymer crosslinking, which produces mechanically robust membranes that are exceptionally effective in inhibiting dendrite growth in a lithium metal battery. The membranes are also reported to enable stable cycling of lithium batteries paired with conventional intercalating cathodes. Our findings appear to provide an important step towards room-temperature dendrite-free batteries. PMID:26634644

The in vivo performance of plasmonic nanobubbles as cell theranostic agents in zebrafish hosting prostate cancer xenografts

PubMed Central

Wagner, Daniel S.; Delk, Nikki A.; Lukianova-Hleb, Ekaterina Y.; Hafner, Jason H.; Farach-Carson, Mary C.; Lapotko, Dmitri O.

2010-01-01

Cell theranostics is a new approach that unites diagnosis, therapy and confirmation (guidance) of the results of therapy in one single process at cell level, thus principally improving both the rapidity and precision of treatment. The ideal theranostic agent will support all three of the above functions in vivo with cellular resolution, allowing individual assessment of disease state and the elimination of diseased cells while leaving healthy cells intact. We have developed and evaluated plasmonic nanobubbles (PNBs) as an in vivo tunable theranostic cellular agent in zebrafish hosting prostate cancer xenografts. PNBs were selectively generated around gold nanoparticles in cancer cells in the zebrafish with short single laser pulses. By varying the energy of the laser pulse, we dynamically tuned the PNB size in a theranostic sequence of two PNBs: an initial small PNB detected a cancer cell through optical scattering, followed by a second bigger PNB, which mechanically ablated this cell without damage to surrounding tissue, while its optical scattering confirmed the destruction of the cell. Thus PNBs supported the diagnosis and guided ablation of individual human cancer cells in a living organism without damage to the host. PMID:20630586

The in vivo performance of plasmonic nanobubbles as cell theranostic agents in zebrafish hosting prostate cancer xenografts.

PubMed

Wagner, Daniel S; Delk, Nikki A; Lukianova-Hleb, Ekaterina Y; Hafner, Jason H; Farach-Carson, Mary C; Lapotko, Dmitri O

2010-10-01

Cell theranostics is a new approach that unites diagnosis, therapy and confirmation (guidance) of the results of therapy in one single process at cell level, thus principally improving both the rapidity and precision of treatment. The ideal theranostic agent will support all three of the above functions in vivo with cellular resolution, allowing individual assessment of disease state and the elimination of diseased cells while leaving healthy cells intact. We have developed and evaluated plasmonic nanobubbles (PNBs) as an in vivo tunable theranostic cellular agent in zebrafish hosting prostate cancer xenografts. PNBs were selectively generated around gold nanoparticles in cancer cells in the zebrafish with short single laser pulses. By varying the energy of the laser pulse, we dynamically tuned the PNB size in a theranostic sequence of two PNBs: an initial small PNB detected a cancer cell through optical scattering, followed by a second bigger PNB, which mechanically ablated this cell without damage to surrounding tissue, while its optical scattering confirmed the destruction of the cell. Thus PNBs supported the diagnosis and guided ablation of individual human cancer cells in a living organism without damage to the host. 2010 Elsevier Ltd. All rights reserved.

Genipin-induced inhibition of uncoupling protein-2 sensitizes drug-resistant cancer cells to cytotoxic agents.

PubMed

Mailloux, Ryan J; Adjeitey, Cyril Nii-Klu; Harper, Mary-Ellen

2010-10-13

Uncoupling protein-2 (UCP2) is known to suppress mitochondrial reactive oxygen species (ROS) production and is employed by drug-resistant cancer cells to mitigate oxidative stress. Using the drug-sensitive HL-60 cells and the drug-resistant MX2 subline as model systems, we show that genipin, a UCP2 inhibitor, sensitizes drug-resistant cells to cytotoxic agents. Increased MX2 cell death was observed upon co-treatment with genipin and different doses of menadione, doxorubicin, and epirubicin. DCFH-DA fluorimetry revealed that the increase in MX2 cell death was accompanied by enhanced cellular ROS levels. The drug-induced increase in ROS was linked to genipin-mediated inhibition of mitochondrial proton leak. State 4 and resting cellular respiratory rates were higher in the MX2 cells in comparison to the HL-60 cells, and the increased respiration was readily suppressed by genipin in the MX2 cells. UCP2 accounted for a remarkable 37% of the resting cellular oxygen consumption indicating that the MX2 cells are functionally reliant on this protein. Higher amounts of UCP2 protein were detected in the MX2 versus the HL-60 mitochondria. The observed effects of genipin were absent in the HL-60 cells pointing to the selectivity of this natural product for drug-resistant cells. The specificity of genipin for UCP2 was confirmed using CHO cells stably expressing UCP2 in which genipin induced an ∼22% decrease in state 4 respiration. These effects were absent in empty vector CHO cells expressing no UCP2. Thus, the chemical inhibition of UCP2 with genipin sensitizes multidrug-resistant cancer cells to cytotoxic agents.

Phytoremediation of TNT: C. roseus hairy roots as a model system

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lauritzen, J.R.; Hughes, J.B.; Shanks, J.V.

Widespread contamination by 2,4,6-trinitrotoluene (TNT) of Soil exists at former munitions production and handling facilities. Phytoremediation may be an effective alternative to existing methods of TNT remediation: incineration is highly expensive and recalcitrant reduction products are formed in composting. Recently, the intrinsic ability of plants to transform TNT has been demonstrated using hairy root cultures of Catharanthus roseus as a model system. Kinetic studies were performed at concentrations of 30 and 50 mg/L TNT in growth medium. The pseudo-first order rate constants for disappearance ranged from 0.0103 to 0.0161 (L/g-day); TNT disappears completely within seven to ten days of exposure.more » The fate of the TNT molecule in plants is also currently under study, mass balance studies were performed with 1-{sup 14}C TNT. After a seven day exposure period, 72% of the label was associated with the roots and 30% was associated with the medium. However, HPLC analysis shows that less than 5% (wt%) of the TNT added is recoverable from both the plants and the media in the form of reduction products. 11 refs., 2 figs.« less

Coupling Deep Transcriptome Analysis with Untargeted Metabolic Profiling in Ophiorrhiza pumila to Further the Understanding of the Biosynthesis of the Anti-Cancer Alkaloid Camptothecin and Anthraquinones

PubMed Central

Yamazaki, Mami; Mochida, Keiichi; Asano, Takashi; Nakabayashi, Ryo; Chiba, Motoaki; Udomson, Nirin; Yamazaki, Yasuyo; Goodenowe, Dayan B.; Sankawa, Ushio; Yoshida, Takuhiro; Toyoda, Atsushi; Totoki, Yasushi; Sakaki, Yoshiyuki; Góngora-Castillo, Elsa; Buell, C. Robin; Sakurai, Tetsuya; Saito, Kazuki

2013-01-01

The Rubiaceae species, Ophiorrhiza pumila, accumulates camptothecin, an anti-cancer alkaloid with a potent DNA topoisomerase I inhibitory activity, as well as anthraquinones that are derived from the combination of the isochorismate and hemiterpenoid pathways. The biosynthesis of these secondary products is active in O. pumila hairy roots yet very low in cell suspension culture. Deep transcriptome analysis was conducted in O. pumila hairy roots and cell suspension cultures using the Illumina platform, yielding a total of 2 Gb of sequence for each sample. We generated a hybrid transcriptome assembly of O. pumila using the Illumina-derived short read sequences and conventional Sanger-derived expressed sequence tag clones derived from a full-length cDNA library constructed using RNA from hairy roots. Among 35,608 non-redundant unigenes, 3,649 were preferentially expressed in hairy roots compared with cell suspension culture. Candidate genes involved in the biosynthetic pathway for the monoterpenoid indole alkaloid camptothecin were identified; specifically, genes involved in post-strictosamide biosynthetic events and genes involved in the biosynthesis of anthraquinones and chlorogenic acid. Untargeted metabolomic analysis by Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) indicated that most of the proposed intermediates in the camptothecin biosynthetic pathway accumulated in hairy roots in a preferential manner compared with cell suspension culture. In addition, a number of anthraquinones and chlorogenic acid preferentially accumulated in hairy roots compared with cell suspension culture. These results suggest that deep transcriptome and metabolome data sets can facilitate the identification of genes and intermediates involved in the biosynthesis of secondary products including camptothecin in O. pumila. PMID:23503598

Downregulation of hPMC2 imparts chemotherapeutic sensitivity to alkylating agents in breast cancer cells.

PubMed

Krishnamurthy, Nirmala; Liu, Lili; Xiong, Xiahui; Zhang, Junran; Montano, Monica M

2015-01-01

Triple negative breast cancer cell lines have been reported to be resistant to the cyotoxic effects of temozolomide (TMZ). We have shown previously that a novel protein, human homolog of Xenopus gene which Prevents Mitotic Catastrophe (hPMC2) has a role in the repair of estrogen-induced abasic sites. Our present study provides evidence that downregulation of hPMC2 in MDA-MB-231 and MDA-MB-468 breast cancer cells treated with temozolomide (TMZ) decreases cell survival. This increased sensitivity to TMZ is associated with an increase in number of apurinic/apyrimidinic (AP) sites in the DNA. We also show that treatment with another alkylating agent, BCNU, results in an increase in AP sites and decrease in cell survival. Quantification of western blot analyses and immunofluorescence experiments reveal that treatment of hPMC2 downregulated cells with TMZ results in an increase in γ-H2AX levels, suggesting an increase in double strand DNA breaks. The enhancement of DNA double strand breaks in TMZ treated cells upon downregulation of hPCM2 is also revealed by the comet assay. Overall, we provide evidence that downregulation of hPMC2 in breast cancer cells increases cytotoxicity of alkylating agents, representing a novel mechanism of treatment for breast cancer. Our data thus has important clinical implications in the management of breast cancer and brings forth potentially new therapeutic strategies.

Target and Non-target Site Mechanisms Developed by Glyphosate-Resistant Hairy beggarticks (Bidens pilosa L.) Populations from Mexico

PubMed Central

Alcántara-de la Cruz, Ricardo; Fernández-Moreno, Pablo T.; Ozuna, Carmen V.; Rojano-Delgado, Antonia M.; Cruz-Hipolito, Hugo E.; Domínguez-Valenzuela, José A.; Barro, Francisco; De Prado, Rafael

2016-01-01

In 2014 hairy beggarticks (Bidens pilosa L.) has been identified as being glyphosate-resistant in citrus orchards from Mexico. The target and non-target site mechanisms involved in the response to glyphosate of two resistant populations (R1 and R2) and one susceptible (S) were studied. Experiments of dose-response, shikimic acid accumulation, uptake-translocation, enzyme activity and 5-enolpyruvyl shikimate-3-phosphate synthase (EPSPS) gene sequencing were carried out in each population. The R1 and R2 populations were 20.4 and 2.8-fold less glyphosate sensitive, respectively, than the S population. The resistant populations showed a lesser shikimic acid accumulation than the S population. In the latter one, 24.9% of 14C-glyphosate was translocated to the roots at 96 h after treatment; in the R1 and R2 populations only 12.9 and 15.5%, respectively, was translocated. Qualitative results confirmed the reduced 14C-glyphosate translocation in the resistant populations. The EPSPS enzyme activity of the S population was 128.4 and 8.5-fold higher than the R1 and R2 populations of glyphosate-treated plants, respectively. A single (Pro-106-Ser), and a double (Thr-102-Ile followed by Pro-106-Ser) mutations were identified in the EPSPS2 gene conferred high resistance in R1 population. Target-site mutations associated with a reduced translocation were responsible for the higher glyphosate resistance in the R1 population. The low-intermediate resistance of the R2 population was mediated by reduced translocation. This is the first glyphosate resistance case confirmed in hairy beggarticks in the world. PMID:27752259

Hydroponics gel as a new electrolyte gelling agent for alkaline zinc-air cells

NASA Astrophysics Data System (ADS)

Othman, R.; Basirun, W. J.; Yahaya, A. H.; Arof, A. K.

The viability of hydroponics gel as a new alkaline electrolyte gelling agent is investigated. Zinc-air cells are fabricated employing 12 wt.% KOH electrolyte immobilised with hydroponics gel. The cells are discharged at constant currents of 5, 50 and 100 mA. XRD and SEM analysis of the anode plates after discharge show that the failure mode is due to the formation of zinc oxide insulating layers and not due to any side reactions between the gel and the plate or the electrolyte.

On-lattice agent-based simulation of populations of cells within the open-source Chaste framework.

PubMed

Figueredo, Grazziela P; Joshi, Tanvi V; Osborne, James M; Byrne, Helen M; Owen, Markus R

2013-04-06

Over the years, agent-based models have been developed that combine cell division and reinforced random walks of cells on a regular lattice, reaction-diffusion equations for nutrients and growth factors; and ordinary differential equations for the subcellular networks regulating the cell cycle. When linked to a vascular layer, this multiple scale model framework has been applied to tumour growth and therapy. Here, we report on the creation of an agent-based multi-scale environment amalgamating the characteristics of these models within a Virtual Physiological Human (VPH) Exemplar Project. This project enables reuse, integration, expansion and sharing of the model and relevant data. The agent-based and reaction-diffusion parts of the multi-scale model have been implemented and are available for download as part of the latest public release of Chaste (Cancer, Heart and Soft Tissue Environment; http://www.cs.ox.ac.uk/chaste/), part of the VPH Toolkit (http://toolkit.vph-noe.eu/). The environment functionalities are verified against the original models, in addition to extra validation of all aspects of the code. In this work, we present the details of the implementation of the agent-based environment, including the system description, the conceptual model, the development of the simulation model and the processes of verification and validation of the simulation results. We explore the potential use of the environment by presenting exemplar applications of the 'what if' scenarios that can easily be studied in the environment. These examples relate to tumour growth, cellular competition for resources and tumour responses to hypoxia (low oxygen levels). We conclude our work by summarizing the future steps for the expansion of the current system.

The Itch-Producing Agents Histamine and Cowhage Activate Separate Populations of Primate Spinothalamic Tract Neurons

PubMed Central

Davidson, Steve; Zhang, Xijing; Yoon, Chul H.; Khasabov, Sergey G.; Simone, Donald A.; Giesler, Glenn J.

2010-01-01

Itch is an everyday sensation, but when associated with disease or infection it can be chronic and debilitating. Several forms of itch can be blocked using antihistamines, but others cannot and these constitute an important clinical problem. Little information is available on the mechanisms underlying itch that is produced by nonhistaminergic mechanisms. We examined the responses of spinothalamic tract neurons to histaminergic and, for the first time, nonhistaminergic forms of itch stimuli. Fifty-seven primate spinothalamic tract (STT) neurons were identified using antidromic activation techniques and examined for their responses to histamine and cowhage, the nonhistaminergic itch-producing spicules covering the pod of the legume Mucuna pruriens. Each examined neuron had a receptive field on the hairy skin of the hindlimb and responded to noxious mechanical stimulation. STT neurons were tested with both pruritogens applied in a random order and we found 12 that responded to histamine and seven to cowhage. Each pruritogen-responsive STT neuron was activated by the chemical algogen capsaicin and two-thirds responded to noxious heat stimuli, demonstrating that these neurons convey chemical, thermal, and mechanical nociceptive information as well. Histamine or cowhage responsive STT neurons were found in both the marginal zone and the deep dorsal horn and were classified as high threshold and wide dynamic range. Unexpectedly, histamine and cowhage never activated the same cell. Our results demonstrate that the spinothalamic tract contains mutually exclusive populations of neurons responsive to histamine or the nonhistaminergic itch-producing agent cowhage. PMID:17855615

Calcium phosphate cement with biofunctional agents and stem cell seeding for dental and craniofacial bone repair.

PubMed

Thein-Han, WahWah; Liu, Jun; Xu, Hockin H K

2012-10-01

Calcium phosphate cement (CPC) can be injected to harden in situ and is promising for dental and craniofacial applications. However, human stem cell attachment to CPC is relatively poor. The objectives of this study were to incorporate biofunctional agents into CPC, and to investigate human umbilical cord mesenchymal stem cell (hUCMSC) seeding on biofunctionalized CPC for osteogenic differentiation for the first time. Five types of biofunctional agents were used: RGD (Arg-Gly-Asp) peptides, human fibronectin (Fn), fibronectin-like engineered polymer protein (FEPP), extracellular matrix Geltrex, and human platelet concentrate. Five biofunctionalized CPC scaffolds were fabricated: CPC-RGD, CPC-Fn, CPC-FEPP, CPC-Geltrex, and CPC-Platelets. The hUCMSC attachment, proliferation, osteogenic differentiation and mineral synthesis were measured. The hUCMSCs on biofunctionalized CPCs had much better cell attachment, proliferation, actin fiber expression, osteogenic differentiation and mineral synthesis, compared to the traditional CPC control. Cell proliferation was increased by an order of magnitude via incorporation of biofunctional agents in CPC (p<0.05). Mineral synthesis on biofunctionalized CPCs was 3-5 folds of those of control (p<0.05). hUCMSCs differentiated with high alkaline phosphatase, Runx2, osteocalcin, and collagen I gene expressions. Mechanical properties of biofunctionalized CPC matched the reported strength and elastic modulus of cancellous bone. A new class of biofunctionalized CPCs was developed, including CPC-RGD, CPC-Fn, CPC-FEPP, CPC-Geltrex, and CPC-Platelets. hUCMSCs on biofunctionalized CPCs had cell density, cell proliferation, actin fiber density, and bone mineralization that were dramatically better than those on traditional CPC. Novel biofunctionalized CPC scaffolds with greatly enhanced human stem cell proliferation and differentiation are promising to facilitate bone regeneration in a wide range of dental, craniofacial and orthopedic

Establishment of Hairy Root Cultures by Agrobacterium Rhizogenes Mediated Transformation of Isatis Tinctoria L. for the Efficient Production of Flavonoids and Evaluation of Antioxidant Activities

PubMed Central

Luo, Meng; Wei, Zuo-Fu; Zu, Yuan-Gang; Ma, Wei; Fu, Yu-Jie

2015-01-01

In this work, Isatis tinctoria hairy root cultures (ITHRCs) were established as an alternative source for flavonoids (FL) production. I. tinctoria hairy root line V was found to be the most efficient line and was further confirmed by the PCR amplification of rolB, rolC and aux1 genes. Culture parameters of ITHRCs were optimized by Box-Behnken design (BBD), and eight bioactive FL constituents (rutin, neohesperidin, buddleoside, liquiritigenin, quercetin, isorhamnetin, kaempferol and isoliquiritigenin) were quali-quantitatively determined by LC-MS/MS. Under optimal conditions, the total FL accumulation of ITHRCs (24 day-old) achieved was 438.10 μg/g dry weight (DW), which exhibited significant superiority as against that of 2 year-old field grown roots (341.73 μg/g DW). Additionally, in vitro antioxidant assays demonstrated that ITHRCs extracts exhibited better antioxidant activities with lower IC50 values (0.41 and 0.39, mg/mL) as compared to those of field grown roots (0.56 and 0.48, mg/mL). To the best of our knowledge, this is the first report describing FL production and antioxidant activities from ITHRCs. PMID:25785699

Establishment of hairy root cultures by Agrobacterium rhizogenes mediated transformation of Isatis tinctoria L. For the efficient production of flavonoids and evaluation of antioxidant activities.

PubMed

Gai, Qing-Yan; Jiao, Jiao; Luo, Meng; Wei, Zuo-Fu; Zu, Yuan-Gang; Ma, Wei; Fu, Yu-Jie

2015-01-01

In this work, Isatis tinctoria hairy root cultures (ITHRCs) were established as an alternative source for flavonoids (FL) production. I. tinctoria hairy root line V was found to be the most efficient line and was further confirmed by the PCR amplification of rolB, rolC and aux1 genes. Culture parameters of ITHRCs were optimized by Box-Behnken design (BBD), and eight bioactive FL constituents (rutin, neohesperidin, buddleoside, liquiritigenin, quercetin, isorhamnetin, kaempferol and isoliquiritigenin) were quali-quantitatively determined by LC-MS/MS. Under optimal conditions, the total FL accumulation of ITHRCs (24 day-old) achieved was 438.10 μg/g dry weight (DW), which exhibited significant superiority as against that of 2 year-old field grown roots (341.73 μg/g DW). Additionally, in vitro antioxidant assays demonstrated that ITHRCs extracts exhibited better antioxidant activities with lower IC₅₀ values (0.41 and 0.39, mg/mL) as compared to those of field grown roots (0.56 and 0.48, mg/mL). To the best of our knowledge, this is the first report describing FL production and antioxidant activities from ITHRCs.

Investigation of Stilbenoids as Potential Therapeutic Agents for Rotavirus Gastroenteritis.

PubMed

Ball, Judith M; Medina-Bolivar, Fabricio; Defrates, Katelyn; Hambleton, Emily; Hurlburt, Megan E; Fang, Lingling; Yang, Tianhong; Nopo-Olazabal, Luis; Atwill, Richard L; Ghai, Pooja; Parr, Rebecca D

2015-01-01

Rotavirus (RV) infections cause severe diarrhea in infants and young children worldwide. Vaccines are available but cost prohibitive for many countries and only reduce severe symptoms. Vaccinated infants continue to shed infectious particles, and studies show decreased efficacy of the RV vaccines in tropical and subtropical countries where they are needed most. Continuing surveillance for new RV strains, assessment of vaccine efficacy, and development of cost effective antiviral drugs remain an important aspect of RV studies. This study was to determine the efficacy of antioxidant and anti-inflammatory stilbenoids to inhibit RV replication. Peanut (A. hypogaea) hairy root cultures were induced to produce stilbenoids, which were purified by high performance countercurrent chromatography (HPCCC) and analyzed by HPLC. HT29.f8 cells were infected with RV in the presence stilbenoids. Cell viability counts showed no cytotoxic effects on HT29.f8 cells. Viral infectivity titers were calculated and comparatively assessed to determine the effects of stilbenoid treatments. Two stilbenoids, trans-arachidin-1 and trans-arachidin-3, show a significant decrease in RV infectivity titers. Western blot analyses performed on the infected cell lysates complemented the infectivity titrations and indicated a significant decrease in viral replication. These studies show the therapeutic potential of the stilbenoids against RV replication.

Investigation of Stilbenoids as Potential Therapeutic Agents for Rotavirus Gastroenteritis

PubMed Central

Ball, Judith M.; Medina-Bolivar, Fabricio; Defrates, Katelyn; Hambleton, Emily; Hurlburt, Megan E.; Fang, Lingling; Yang, Tianhong; Nopo-Olazabal, Luis; Atwill, Richard L.; Ghai, Pooja; Parr, Rebecca D.

2015-01-01

Rotavirus (RV) infections cause severe diarrhea in infants and young children worldwide. Vaccines are available but cost prohibitive for many countries and only reduce severe symptoms. Vaccinated infants continue to shed infectious particles, and studies show decreased efficacy of the RV vaccines in tropical and subtropical countries where they are needed most. Continuing surveillance for new RV strains, assessment of vaccine efficacy, and development of cost effective antiviral drugs remain an important aspect of RV studies. This study was to determine the efficacy of antioxidant and anti-inflammatory stilbenoids to inhibit RV replication. Peanut (A. hypogaea) hairy root cultures were induced to produce stilbenoids, which were purified by high performance countercurrent chromatography (HPCCC) and analyzed by HPLC. HT29.f8 cells were infected with RV in the presence stilbenoids. Cell viability counts showed no cytotoxic effects on HT29.f8 cells. Viral infectivity titers were calculated and comparatively assessed to determine the effects of stilbenoid treatments. Two stilbenoids, trans-arachidin-1 and trans-arachidin-3, show a significant decrease in RV infectivity titers. Western blot analyses performed on the infected cell lysates complemented the infectivity titrations and indicated a significant decrease in viral replication. These studies show the therapeutic potential of the stilbenoids against RV replication. PMID:26379708

Lemongrass essential oil and citral inhibit Src/Stat3 activity and suppress the proliferation/survival of small-cell lung cancer cells, alone or in combination with chemotherapeutic agents.

PubMed

Maruoka, Takayuki; Kitanaka, Akira; Kubota, Yoshitsugu; Yamaoka, Genji; Kameda, Tomohiro; Imataki, Osamu; Dobashi, Hiroaki; Bandoh, Shuji; Kadowaki, Norimitsu; Tanaka, Terukazu

2018-03-13

Small-cell lung cancer (SCLC) is intractable due to its high propensity for relapse. Novel agents are thus needed for SCLC treatment. Lemongrass essential oil (LG-EO) and its major constituent, citral, have been reported to inhibit the proliferation and survival of several types of cancer cells. However, the precise mechanisms through which LG-EO and citral exert their effects on SCLC cells have not been fully elucidated. SCLC cells express Src and have high levels of Src-tyrosine kinase (Src-TK) activity. In most SCLC cell lines, constitutive phosphorylation of Stat3(Y705), which is essential for its activation, has been detected. Src-TK can phosphorylate Stat3(Y705), and activated Stat3 promotes the expression of the anti-apoptotic factors Bcl-xL and Mcl-1. In the present study, LG-EO and citral prevented Src-TK from phosphorylating Stat3(Y705), resulting in decreased Bcl-xL and Mcl-1 expression, in turn suppressing the proliferation/survival of SCLC cells. To confirm these findings, the wild-type-src gene was transfected into the LU135 SCLC cell line (LU135‑wt-src), in which Src and activated phospho-Stat3(Y705) were overexpressed. The suppression of cell proliferation and the induction of apoptosis by treatment with LG-EO or citral were significantly attenuated in the LU135-wt-src cells compared with the control LU135-mock cells. The signal transducer and activator of transcription 3 (Stat3) signaling pathway is also associated with intrinsic drug resistance. LU135-wt-src cells were significantly resistant to conventional chemotherapeutic agents compared with LU135-mock cells. The combined effects of citral and each conventional chemotherapeutic agent on SCLC cells were also evaluated. The combination treatment exerted additive or more prominent effects on LU135-wt-src, LU165 and MN1112 cells, which are relatively chemoresistant SCLC cells. These findings suggest that either LG-EO or citral, alone or in combination with chemotherapeutic agents, may be a

Isolation and structure elucidation of a new indole alkaloid from Rauvolfia serpentina hairy root culture: the first naturally occurring alkaloid of the raumacline group.

PubMed

Sheludko, Yuri; Gerasimenko, Irina; Kolshorn, Heinz; Stöckigt, Joachim

2002-05-01

A new monoterpenoid indole alkaloid, 10-hydroxy- N(alpha)-demethyl-19,20-dehydroraumacline ( 1), was isolated as a mixture of E- and Z-isomers from hairy root culture of Rauvolfia serpentina Benth. ex Kurz (Apocynaceae) and the structure was determined by 1D and 2D NMR analyses. The new indole alkaloid represents the first naturally occurring alkaloid of the raumacline group and its putative biosynthetical pathway is discussed.

Targeting neddylation induces DNA damage and checkpoint activation and sensitizes chronic lymphocytic leukemia B cells to alkylating agents.

PubMed

Paiva, C; Godbersen, J C; Berger, A; Brown, J R; Danilov, A V

2015-07-09

Microenvironment-mediated upregulation of the B-cell receptor (BCR) and nuclear factor-κB (NF-κB) signaling in CLL cells resident in the lymph node and bone marrow promotes apoptosis evasion and clonal expansion. We recently reported that MLN4924 (pevonedistat), an investigational agent that inhibits the NEDD8-activating enzyme (NAE), abrogates stromal-mediated NF-κB pathway activity and CLL cell survival. However, the NAE pathway also assists degradation of multiple other substrates. MLN4924 has been shown to induce DNA damage and cell cycle arrest, but the importance of this mechanism in primary neoplastic B cells has not been studied. Here we mimicked the lymph node microenvironment using CD40 ligand (CD40L)-expressing stroma and interleukin-21 (IL-21) to find that inducing proliferation of the primary CLL cells conferred enhanced sensitivity to NAE inhibition. Treatment of the CD40-stimulated CLL cells with MLN4924 resulted in deregulation of Cdt1, a DNA replication licensing factor, and cell cycle inhibitors p21 and p27. This led to DNA damage, checkpoint activation and G2 arrest. Alkylating agents bendamustine and chlorambucil enhanced MLN4924-mediated DNA damage and apoptosis. These events were more prominent in cells stimulated with IL-21 compared with CD40L alone, indicating that, following NAE inhibition, the culture conditions were able to direct CLL cell fate from an NF-κB inhibition to a Cdt1 induction program. Our data provide insight into the biological consequences of targeting NAE in CLL and serves as further rationale for studying the clinical activity of MLN4924 in CLL, particularly in combination with alkylating agents.

Targeting neddylation induces DNA damage and checkpoint activation and sensitizes chronic lymphocytic leukemia B cells to alkylating agents

PubMed Central

Paiva, C; Godbersen, J C; Berger, A; Brown, J R; Danilov, A V

2015-01-01

Microenvironment-mediated upregulation of the B-cell receptor (BCR) and nuclear factor-κB (NF-κB) signaling in CLL cells resident in the lymph node and bone marrow promotes apoptosis evasion and clonal expansion. We recently reported that MLN4924 (pevonedistat), an investigational agent that inhibits the NEDD8-activating enzyme (NAE), abrogates stromal-mediated NF-κB pathway activity and CLL cell survival. However, the NAE pathway also assists degradation of multiple other substrates. MLN4924 has been shown to induce DNA damage and cell cycle arrest, but the importance of this mechanism in primary neoplastic B cells has not been studied. Here we mimicked the lymph node microenvironment using CD40 ligand (CD40L)-expressing stroma and interleukin-21 (IL-21) to find that inducing proliferation of the primary CLL cells conferred enhanced sensitivity to NAE inhibition. Treatment of the CD40-stimulated CLL cells with MLN4924 resulted in deregulation of Cdt1, a DNA replication licensing factor, and cell cycle inhibitors p21 and p27. This led to DNA damage, checkpoint activation and G2 arrest. Alkylating agents bendamustine and chlorambucil enhanced MLN4924-mediated DNA damage and apoptosis. These events were more prominent in cells stimulated with IL-21 compared with CD40L alone, indicating that, following NAE inhibition, the culture conditions were able to direct CLL cell fate from an NF-κB inhibition to a Cdt1 induction program. Our data provide insight into the biological consequences of targeting NAE in CLL and serves as further rationale for studying the clinical activity of MLN4924 in CLL, particularly in combination with alkylating agents. PMID:26158513

[The effect of sodium phenylbutyrate to agents used in induction chemotherapy on laryngeal carcinoma cells Hep-2 in vitro].

PubMed

Gao, Jing; Ruan, Xinyong; Pan, Xinliang; Xu, Fenglei; Lei, Dapeng; Liu, Dayu

2005-08-01

To study the effect of sodium phenylbutyrate when it combined with agents used in induction chemotherapy on laryngeal carcinoma cells Hep-2 in vitro. MTT were used to examine the growth inhibition of Hep-2 cells treated by the combination of PB with 5-FU or CDDP in vitro. When 5-FU or CDDP combined with PB respectively, there was significantly difference between every two dose groups of the two agents or every dose group and control group ( P < 0.05). When the dosage of 5-FU or CDDP was definition,there was significantly difference between every two dose groups of PB ( P < 0.05). PB could enhance the cytotoxic effects of agents used in induction chemotherapy on laryngeal carcinoma cells Hep-2 in vitro, which showed the possibility in reinforcement the treatment effect and reduction the occurrence of the complication and toxic reaction of induction chemotherapy on laryngeal carcinoma.

Cytotoxic and apoptotic effects of bortezomib and gefitinib compared to alkylating agents on human glioblastoma cells.

PubMed

Pédeboscq, Stéphane; L'Azou, Béatrice; Passagne, Isabelle; De Giorgi, Francesca; Ichas, François; Pometan, Jean-Paul; Cambar, Jean

2008-01-01

Glioblastoma is a malignant astrocytic tumor with a median survival of about 12 months for which new therapeutic strategies are required. We therefore examined the cytotoxicity of anticancer drugs with different mechanisms of action on two human glioblastoma cell lines expressing various levels of EGFR (epidermal growth factor receptor). Apoptosis induced by these anticancer agents was evaluated by flow cytometry. The cytotoxicity of alkylating drugs followed a dose-effect curve and cytotoxicity index values were lower with carboplatin than with BCNU and temozolomide. Anti-EGFR gefitinib (10 microM) cytotoxicity on DBTRG.05-MG expressing high levels of EGFR was significantly higher than on U87-MG expressing low levels of EGFR. Carboplatin and temozolomide cytotoxicity was potentiated with the addition of gefitinib on DBTRG.05-MG. Among the anticancer agents tested, the proteasome inhibitor bortezomib was the most cytotoxic with very low IC50 on the two cell lines. Moreover, all anticancer drugs tested induced apoptosis in a concentration-dependent manner. Bortezomib proved to be a more potent inductor of apoptosis than gefitinib and alkylating agents. These results show the efficacy of bortezomib and of the association between conventional chemotherapy and gefitinib on glioblastoma cells and therefore suggest the interest of these molecules in the treatment of glioblastoma.

Cell fusing agent virus and dengue virus mutually interact in Aedes aegypti cell lines.

PubMed

Zhang, Guangmei; Asad, Sultan; Khromykh, Alexander A; Asgari, Sassan

2017-07-31

The genus Flavivirus contains more than 70 single-stranded, positive-sense arthropod-borne RNA viruses. Some flaviviruses are particularly medically important to humans and other vertebrates including dengue virus (DENV), West Nile virus, and yellow fever virus. These viruses are transmitted to vertebrates by mosquitoes and other arthropod species. Mosquitoes are also infected by insect-specific flaviviruses (ISFs) that do not appear to be infective to vertebrates. Cell fusing agent virus (CFAV) was the first described ISF, which was discovered in an Aedes aegypti cell culture. We found that while CFAV infection could be significantly reduced by application of RNAi against the NS5 gene, removal of the treatment led to quick restoration of CFAV replication. Interestingly, we found that CFAV infection significantly enhanced replication of DENV, and vice versa, DENV infection significantly enhanced replication of CFAV in mosquito cells. We have shown that CFAV infection leads to increase in the expression of ribonuclease kappa (RNASEK), which is known to promote infection of viruses that rely on endocytosis and pH-dependent entry. Knockdown of RNASEK by dsRNA resulted in reduced DENV replication. Thus, increased expression of RNASEK induced by CFAV is likely to contribute to enhanced DENV replication in CFAV-infected cells.

Biomorphic Multi-Agent Architecture for Persistent Computing

NASA Technical Reports Server (NTRS)

Lodding, Kenneth N.; Brewster, Paul

2009-01-01

A multi-agent software/hardware architecture, inspired by the multicellular nature of living organisms, has been proposed as the basis of design of a robust, reliable, persistent computing system. Just as a multicellular organism can adapt to changing environmental conditions and can survive despite the failure of individual cells, a multi-agent computing system, as envisioned, could adapt to changing hardware, software, and environmental conditions. In particular, the computing system could continue to function (perhaps at a reduced but still reasonable level of performance) if one or more component( s) of the system were to fail. One of the defining characteristics of a multicellular organism is unity of purpose. In biology, the purpose is survival of the organism. The purpose of the proposed multi-agent architecture is to provide a persistent computing environment in harsh conditions in which repair is difficult or impossible. A multi-agent, organism-like computing system would be a single entity built from agents or cells. Each agent or cell would be a discrete hardware processing unit that would include a data processor with local memory, an internal clock, and a suite of communication equipment capable of both local line-of-sight communications and global broadcast communications. Some cells, denoted specialist cells, could contain such additional hardware as sensors and emitters. Each cell would be independent in the sense that there would be no global clock, no global (shared) memory, no pre-assigned cell identifiers, no pre-defined network topology, and no centralized brain or control structure. Like each cell in a living organism, each agent or cell of the computing system would contain a full description of the system encoded as genes, but in this case, the genes would be components of a software genome.

Targeting RNA polymerase I transcription machinery in cancer cells by a novel monofunctional platinum-based agent.

PubMed

Zhang, Zhen-Lei; Zhao, Chun-Lai; Chen, Qian; Xu, Kai; Qiao, Xin; Xu, Jing-Yuan

2018-06-01

Aberrant ribosome biogenesis and enlarged nucleoli have long been used by pathologists as a marker of aggressive tumors. Suppression of RNA polymerase I (Pol I) transcription machinery within the nucleolus could be a direct way to trigger the nucleolar stress and to inhibit the rapid proliferation of cancer cells. Here we modified cisplatin with an analogue of the selective inhibitor of RNA polymerase I-mediated transcription BMH-21 to develop a novel platinum-based Pol I selective inhibitor. We show that this novel monofunctional platinum-based agent, P1-B1, had enhanced antitumor activity of up to 17-fold greater than the clinical drug cisplatin in cisplatin-resistant non-small cell lung cancer cells. P1-B1 also had significantly lower cytotoxicity compared to cisplatin as well as the Pol I selective inhibitor BMH-21 in MRC-5 normal lung fibroblast cells, and the selectivity index (SI) greatly increases. Mechanistic investigations revealed that P1-B1 displayed significant nucleolar accumulation, selectively inhibited Pol I transcription, and induced nucleolar stress, leading to S-phase arrest and apoptosis. Our results suggest that the effects of P1-B1 are mechanistically distinct from those of conventional platinum agents and the recently described non-classical platinum compounds and that functionalizing platinum-based agents with directly Pol I transcription inhibition properties may represent an improved modality for cancer treatment. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Cell Death Pathways and Phthalocyanine as an Efficient Agent for Photodynamic Cancer Therapy

PubMed Central

Mfouo-Tynga, Ivan; Abrahamse, Heidi

2015-01-01

The mechanisms of cell death can be predetermined (programmed) or not and categorized into apoptotic, autophagic and necrotic pathways. The process of Hayflick limits completes the execution of death-related mechanisms. Reactive oxygen species (ROS) are associated with oxidative stress and subsequent cytodamage by oxidizing and degrading cell components. ROS are also involved in immune responses, where they stabilize and activate both hypoxia-inducible factors and phagocytic effectors. ROS production and presence enhance cytodamage and photodynamic-induced cell death. Photodynamic cancer therapy (PDT) uses non-toxic chemotherapeutic agents, photosensitizer (PS), to initiate a light-dependent and ROS-related cell death. Phthalocyanines (PCs) are third generation and stable PSs with improved photochemical abilities. They are effective inducers of cell death in various neoplastic models. The metallated PCs localize in critical cellular organelles and are better inducers of cell death than other previous generation PSs as they favor mainly apoptotic cell death events. PMID:25955645

Camptothecin-loaded fusogenic nanodroplets as ultrasound theranostic agent in stem cell-mediated drug-delivery system.

PubMed

Ho, Yi-Ju; Chiang, Yu-Jung; Kang, Shih-Tsung; Fan, Ching-Hsiang; Yeh, Chih-Kuang

2018-05-28

Adipose-derived stem cells (ADSCs) have been utilized in cellular delivery systems to carry therapeutic agents into tumors by migration. Drug-loaded nanodroplets release drugs and form bubbles after acoustic droplet vaporization (ADV) triggered by ultrasound stimulation, providing a system for ultrasound-induced cellular delivery of theranostic agents. In order to improve the efficiency of drug release, fusogenic nanodroplets were designed to go from nano to micron size upon uptake by ADSCs for reducing ADV threshold. The purpose of our study was to demonstrate the utility of camptothecin-loaded fusogenic nanodroplets (CPT-FNDs) as ultrasound theranostic agents in an ADSCs delivery system. CPT-FNDs showed an increase in size from 81.6 ± 3.5 to 1043.5 ± 28.3 nm and improved CPT release from 22.0 ± 1.8% to 37.6 ± 2.1%, demonstrating the fusion ability of CPT-FNDs. CPT-FNDs-loaded ADSCs demonstrated a cell viability of 77 ± 4%, and the in vitro migration ability was 3.2 ± 1.2-fold for the tumor condition compared to the cell growth condition. Ultrasound enhancement imaging showed intratumoral ADV-generated bubble formation (increasing 3.24 ± 0.47 dB) triggered by ultrasound after CPT-FNDs-loaded ADSCs migration into B16F0 tumors. Histological images revealed intratumoral distribution of CPT-FNDs-loaded ADSCs and tissue damage due to the ADV. The CPT-FNDs can be used as theranostic agents in an ADSCs delivery system to provide the ultrasound contrast imaging and deliver combination therapy of drug release and physical damage after ADV. Copyright © 2018 Elsevier B.V. All rights reserved.

Phylogeography of screaming hairy armadillo Chaetophractus vellerosus: Successive disjunctions and extinctions due to cyclical climatic changes in southern South America.

PubMed

Poljak, Sebastián; Ferreiro, Alejandro M; Chiappero, Marina B; Sánchez, Julieta; Gabrielli, Magalí; Lizarralde, Marta S

2018-01-01

Little is known about phylogeography of armadillo species native to southern South America. In this study we describe the phylogeography of the screaming hairy armadillo Chaetophractus vellerosus, discuss previous hypothesis about the origin of its disjunct distribution and propose an alternative one, based on novel information on genetic variability. Variation of partial sequences of mitochondrial DNA Control Region (CR) from 73 individuals from 23 localities were analyzed to carry out a phylogeographic analysis using neutrality tests, mismatch distribution, median-joining (MJ) network and paleontological records. We found 17 polymorphic sites resulting in 15 haplotypes. Two new geographic records that expand known distribution of the species are presented; one of them links the distributions of recently synonimized species C. nationi and C. vellerosus. Screaming hairy armadillo phylogeographic pattern can be addressed as category V of Avise: common widespread linages plus closely related lineages confined to one or a few nearby locales each. The older linages are distributed in the north-central area of the species distribution range in Argentina (i.e. ancestral area of distribution). C. vellerosus seems to be a low vagility species that expanded, and probably is expanding, its distribution range while presents signs of genetic structuring in central areas. To explain the disjunct distribution, a hypothesis of extinction of the species in intermediate areas due to quaternary climatic shift to more humid conditions was proposed. We offer an alternative explanation: long distance colonization, based on null genetic variability, paleontological record and evidence of alternance of cold/arid and temperate/humid climatic periods during the last million years in southern South America.

Quantitative Analyses of Synergistic Responses between Cannabidiol and DNA-Damaging Agents on the Proliferation and Viability of Glioblastoma and Neural Progenitor Cells in Culture

PubMed Central

Deng, Liting; Ng, Lindsay; Ozawa, Tatsuya

2017-01-01

Evidence suggests that the nonpsychotropic cannabis-derived compound, cannabidiol (CBD), has antineoplastic activity in multiple types of cancers, including glioblastoma multiforme (GBM). DNA-damaging agents remain the main standard of care treatment available for patients diagnosed with GBM. Here we studied the antiproliferative and cell-killing activity of CBD alone and in combination with DNA-damaging agents (temozolomide, carmustine, or cisplatin) in several human GBM cell lines and in mouse primary GBM cells in cultures. This activity was also studied in mouse neural progenitor cells (NPCs) in culture to assess for potential central nervous system toxicity. We found that CBD induced a dose-dependent reduction of both proliferation and viability of all cells with similar potencies, suggesting no preferential activity for cancer cells. Hill plot analysis indicates an allosteric mechanism of action triggered by CBD in all cells. Cotreatment regimens combining CBD and DNA-damaging agents produced synergistic antiproliferating and cell-killing responses over a limited range of concentrations in all human GBM cell lines and mouse GBM cells as well as in mouse NPCs. Remarkably, antagonistic responses occurred at low concentrations in select human GBM cell lines and in mouse GBM cells. Our study suggests limited synergistic activity when combining CBD and DNA-damaging agents in treating GBM cells, along with little to no therapeutic window when considering NPCs. PMID:27821713

Fluorine-19 MRI Contrast Agents for Cell Tracking and Lung Imaging

PubMed Central

Fox, Matthew S.; Gaudet, Jeffrey M.; Foster, Paula J.

2015-01-01

Fluorine-19 (19F)-based contrast agents for magnetic resonance imaging stand to revolutionize imaging-based research and clinical trials in several fields of medical intervention. First, their use in characterizing in vivo cell behavior may help bring cellular therapy closer to clinical acceptance. Second, their use in lung imaging provides novel noninvasive interrogation of the ventilated airspaces without the need for complicated, hard-to-distribute hardware. This article reviews the current state of 19F-based cell tracking and lung imaging using magnetic resonance imaging and describes the link between the methods across these fields and how they may mutually benefit from solutions to mutual problems encountered when imaging 19F-containing compounds, as well as hardware and software advancements. PMID:27042089

CIP-36, a novel topoisomerase II-targeting agent, induces the apoptosis of multidrug-resistant cancer cells in vitro.

PubMed

Cao, Bo; Chen, Hong; Gao, Ying; Niu, Cong; Zhang, Yuan; Li, Ling

2015-03-01

The need to overcome cancer multidrug resistance (MDR) has fueled considerable interest in the development of novel synthetic antitumor agents with cytotoxicity against cancer cell lines with MDR. In this study, we aimed to investigate CIP-36, a novel podophyllotoxin derivative, for its inhibitory effects on human cancer cells from multiple sources, particularly cells with MDR in vitro. The human leukemia cell line, K562, and the adriamycin-resistant subline, K562/A02, were exposed to CIP-36 or anticancer agents, and various morphological and biochemical properties were assessed by Hoechst 33342 staining under a fluorescence microscope. Subsequently, cytotoxicity, cell growth curves and the cell cycle were analyzed. Finally, the effects of CIP-36 on topoisomerase IIα (Topo IIα) activity were determined. Treatment with CIP-36 significantly inhibited the growth of the K562 and MDR K562/A02 cells. Our data demonstrated that CIP-36 induced apoptosis, inhibited cell cycle progression and inhibited Topo IIα activity. These findings suggest that CIP-36 has the potential to overcome the multidrug resistance of K562/A02 cells by mediating Topo IIα activity.

Tailoring tobacco hairy root metabolism for the production of stilbenes.

PubMed

Hidalgo, Diego; Georgiev, Milen; Marchev, Andrey; Bru-Martínez, Roque; Cusido, Rosa M; Corchete, Purificación; Palazon, Javier

2017-12-21

Tobacco hairy root (HR) cultures, which have been widely used for the heterologous production of target compounds, have an innate capacity to bioconvert exogenous t-resveratrol (t-R) into t-piceatannol (t-Pn) and t-pterostilbene (t-Pt). We established genetically engineered HR carrying the gene encoding stilbene synthase (STS) from Vitis vinifera and/or the transcription factor (TF) AtMYB12 from Arabidopsis thaliana, in order to generate a holistic response in the phenylpropanoid pathway and coordinate the up-regulation of multiple metabolic steps. Additionally, an artificial microRNA for chalcone synthase (amiRNA CHS) was utilized to arrest the normal flux through the endogenous chalcone synthase (CHS) enzyme, which would otherwise compete for precursors with the STS enzyme imported for the flux deviation. The transgenic HR were able to biosynthesize the target stilbenes, achieving a production of 40 μg L -1 of t-R, which was partially metabolized into t-Pn and t-Pt (up to 2.2 μg L -1 and 86.4 μg L -1 , respectively), as well as its glucoside piceid (up to 339.7 μg L -1 ). Major metabolic perturbations were caused by the TF AtMYB12, affecting both primary and secondary metabolism, which confirms the complexity of biotechnological systems based on seed plant in vitro cultures for the heterologous production of high-value molecules.

Identification of agents effective against multiple toxins and viruses by host-oriented cell targeting.

PubMed

Zilbermintz, Leeor; Leonardi, William; Jeong, Sun-Young; Sjodt, Megan; McComb, Ryan; Ho, Chi-Lee C; Retterer, Cary; Gharaibeh, Dima; Zamani, Rouzbeh; Soloveva, Veronica; Bavari, Sina; Levitin, Anastasia; West, Joel; Bradley, Kenneth A; Clubb, Robert T; Cohen, Stanley N; Gupta, Vivek; Martchenko, Mikhail

2015-08-27

A longstanding and still-increasing threat to the effective treatment of infectious diseases is resistance to antimicrobial countermeasures. Potentially, the targeting of host proteins and pathways essential for the detrimental effects of pathogens offers an approach that may discover broad-spectrum anti-pathogen countermeasures and circumvent the effects of pathogen mutations leading to resistance. Here we report implementation of a strategy for discovering broad-spectrum host-oriented therapies against multiple pathogenic agents by multiplex screening of drugs for protection against the detrimental effects of multiple pathogens, identification of host cell pathways inhibited by the drug, and screening for effects of the agent on other pathogens exploiting the same pathway. We show that a clinically used antimalarial drug, Amodiaquine, discovered by this strategy, protects host cells against infection by multiple toxins and viruses by inhibiting host cathepsin B. Our results reveal the practicality of discovering broadly acting anti-pathogen countermeasures that target host proteins exploited by pathogens.

APC10.1 cells as a model for assessing the efficacy of potential chemopreventive agents in the Apc(Min) mouse model in vivo.

PubMed

Sale, Stewart; Fong, Isabel L; de Giovanni, Carla; Landuzzi, Lorena; Brown, Karen; Steward, William P; Gescher, Andreas J

2009-11-01

Apc(Min) mice are widely used for mechanism and efficacy studies associated with the development of chemopreventive agents. APC10.1 cells have been derived from Apc(Min) mouse adenomas and retain the heterozygous Apc genotype. We tested the hypothesis that this cell type may provide an in vitro model to predict chemopreventive activity of agents in the Apc(Min) mouse in vivo. The growth inhibitory properties of 14 putative colorectal cancer chemopreventive agents, tricin, apigenin, 3',4',5',5,7-pentamethoxyflavone, resveratrol, curcumin, 3,4-methylenedioxy-3',4',5'-trimethoxychalcone (DMU135), 3,4,5,4'-tetramethoxystilbene (DMU212), celecoxib, aspirin, piroxicam, all-trans-retinoic acid, difluoromethylornithine (DFMO), quercetin and cyanidin-3-glucoside, were studied in this cell line, and the IC(50) values were calculated. The IC(50) values were plotted against previously published data of reduction of adenoma numbers caused by these agents in Apc(Min) mice. The correlation co-efficient was 0.678 (p<0.01), suggesting that there was a tentative correlation between the ability to inhibit the growth of APC10.1 cells and the ability to delay adenoma development in vivo. If this relationship is supported by using further agents, APC10.1 cells may serve in the future as an initial screen to prioritise compounds for assessing chemopreventive efficacy in Apc(Min) mice in vivo. Such a screen could reduce the number of animals required to find active agents, help reduce costs and increase throughput.

The epigenetic agents suberoylanilide hydroxamic acid and 5‑AZA‑2' deoxycytidine decrease cell proliferation, induce cell death and delay the growth of MiaPaCa2 pancreatic cancer cells in vivo.

PubMed

Susanto, Johana M; Colvin, Emily K; Pinese, Mark; Chang, David K; Pajic, Marina; Mawson, Amanda; Caldon, C Elizabeth; Musgrove, Elizabeth A; Henshall, Susan M; Sutherland, Robert L; Biankin, Andrew V; Scarlett, Christopher J

2015-05-01

Despite incremental advances in the diagnosis and treatment for pancreatic cancer (PC), the 5‑year survival rate remains <5%. Novel therapies to increase survival and quality of life for PC patients are desperately needed. Epigenetic thera-peutic agents such as histone deacetylase inhibitors (HDACi) and DNA methyltransferase inhibitors (DNMTi) have demonstrated therapeutic benefits in human cancer. We assessed the efficacy of these epigenetic therapeutic agents as potential therapies for PC using in vitro and in vivo models. Treatment with HDACi [suberoylanilide hydroxamic acid (SAHA)] and DNMTi [5‑AZA‑2' deoxycytidine (5‑AZA‑dc)] decreased cell proliferation in MiaPaCa2 cells, and SAHA treatment, with or without 5‑AZA‑dc, resulted in higher cell death and lower DNA synthesis compared to 5‑AZA‑dc alone and controls (DMSO). Further, combination treatment with SAHA and 5‑AZA‑dc significantly increased expression of p21WAF1, leading to G1 arrest. Treatment with epigenetic agents delayed tumour growth in vivo, but did not decrease growth of established pancreatic tumours. In conclusion, these data demonstrate a potential role for epigenetic modifier drugs for the management of PC, specifically in the chemoprevention of PC, in combination with other chemotherapeutic agents.

Genetically engineered hairy root cultures of Hyoscyamus senecionis and H. muticus: ploidy as a promising parameter in the metabolic engineering of tropane alkaloids.

PubMed

Dehghan, Esmaeil; Reed, Darwin W; Covello, Patrick S; Hasanpour, Zeinab; Palazon, Javier; Oksman-Caldentey, Kirsi-Marja; Ahmadi, Farajollah Shahriari

2017-10-01

Tetraploidy improves overexpression of h6h and scopolamine production of H. muticus, while in H. senecionis, pmt overexpression and elicitation can be used as effective methods for increasing tropane alkaloids. The effects of metabolic engineering in a polyploid context were studied by overexpression of h6h in the tetraploid hairy root cultures of H. muticus. Flow cytometry analysis indicated genetic stability in the majority of the clones, while only a few clones showed genetic instability. Among all the diploid and tetraploid clones, the highest level of h6h transgene expression and scopolamine accumulation was interestingly observed in the tetraploid clones of H. muticus. Therefore, metabolic engineering of the tropane biosynthetic pathway in polyploids is suggested as a potential system for increasing the production of tropane alkaloids. Transgenic hairy root cultures of Hyoscyamus senecionis were also established. While overexpression of pmt in H. senecionis was correlated with a sharp increase in hyoscyamine production, the h6h-overexpressing clones were not able to accumulate higher levels of scopolamine than the leaves of intact plants. Applying methyl jasmonate was followed by a sharp increase in the expression of pmt and a drop in the expression of tropinone reductase II (trII) which consequently resulted in the higher biosynthesis of hyoscyamine and total alkaloids in H. senecionis.

Induction of brain tumor stem cell apoptosis by FTY720: a potential therapeutic agent for glioblastoma.

PubMed

Estrada-Bernal, Adriana; Palanichamy, Kamalakannan; Ray Chaudhury, Abhik; Van Brocklyn, James R

2012-04-01

FTY720 is a sphingosine analogue that down regulates expression of sphingosine-1-phosphate receptors and causes apoptosis of multiple tumor cell types, including glioma cells. This study examined the effect of FTY720 on brain tumor stem cells (BTSCs) derived from human glioblastoma (GBM) tissue. FTY720 treatment of BTSCs led to rapid inactivation of ERK MAP kinase, leading to upregulation of the BH3-only protein Bim and apoptosis. In combination with temozolomide (TMZ), the current standard chemotherapeutic agent for GBM, FTY720 synergistically induced BTSC apoptosis. FTY720 also slowed growth of intracranial xenograft tumors in nude mice and augmented the therapeutic effect of TMZ, leading to enhanced survival. Furthermore, the combination of FTY720 and TMZ decreased the invasiveness of BTSCs in mouse brains. FTY720 is known to cross the blood-brain barrier and recently received Food and Drug Administration approval for treatment of relapsing multiple sclerosis. Thus, FTY720 is an excellent potential therapeutic agent for treatment of GBM.

Protection against radiation-induced oxidative stress in cultured human epithelial cells by treatment with antioxidant agents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wan, X. Steven; Ware, Jeffrey H.; Zhou, Zhaozong

2006-04-01

Purpose: To evaluate the protective effects of antioxidant agents against space radiation-induced oxidative stress in cultured human epithelial cells. Methods and Materials: The effects of selected concentrations of N-acetylcysteine, ascorbic acid, sodium ascorbate, co-enzyme Q10, {alpha}-lipoic acid, L-selenomethionine, and vitamin E succinate on radiation-induced oxidative stress were evaluated in MCF10 human breast epithelial cells exposed to radiation with X-rays, {gamma}-rays, protons, or high mass, high atomic number, and high energy particles using a dichlorofluorescein assay. Results: The results demonstrated that these antioxidants are effective in protecting against radiation-induced oxidative stress and complete or nearly complete protection was achieved by treatingmore » the cells with a combination of these agents before and during the radiation exposure. Conclusion: The combination of antioxidants evaluated in this study is likely be a promising countermeasure for protection against space radiation-induced adverse biologic effects.« less

[Supramolecular Agents for Theranostics].

PubMed

Deyev, S M; Lebedenko, E N

2015-01-01

This mini-review summarizes recent data obtained in the process of creation of a versatile module platform suitable for construction of supramolecular theranostic agents. As an example, we consider multifunctional hybrid agents for imaging and elimination of cancer cells. The use of an adapter protein system barnase:barstar for producing targeted multifunctional hybrid structures on the basis of highly specific peptides and mini-antibodies as addressing modules and recombinant proteins and/or nanoparticles of different nature (quantum dots, nanogold, magnetic nanoparticles, nanodiamonds, upconverting nanophosphores, polymer nanoparticles) as agents visualizing and damaging cancer cells is described. New perspectives for creation of selective and highly effective compounds for theranostics and personified medicine are contemplated.

Rituximab, Romidepsin, and Lenalidomide in Treating Patients With Recurrent or Refractory B-cell Non-Hodgkin Lymphoma

ClinicalTrials.gov

2016-08-09

B-cell Adult Acute Lymphoblastic Leukemia; Extranodal Marginal Zone B-cell Lymphoma of Mucosa-associated Lymphoid Tissue; Intraocular Lymphoma; Nodal Marginal Zone B-cell Lymphoma; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Adult Burkitt Lymphoma; Recurrent Adult Diffuse Large Cell Lymphoma; Recurrent Adult Diffuse Mixed Cell Lymphoma; Recurrent Adult Diffuse Small Cleaved Cell Lymphoma; Recurrent Adult Grade III Lymphomatoid Granulomatosis; Recurrent Adult Immunoblastic Large Cell Lymphoma; Recurrent Adult Lymphoblastic Lymphoma; Recurrent Grade 1 Follicular Lymphoma; Recurrent Grade 2 Follicular Lymphoma; Recurrent Grade 3 Follicular Lymphoma; Recurrent Mantle Cell Lymphoma; Recurrent Marginal Zone Lymphoma; Recurrent Small Lymphocytic Lymphoma; Refractory Hairy Cell Leukemia; Small Intestine Lymphoma; Splenic Marginal Zone Lymphoma; Testicular Lymphoma; Waldenström Macroglobulinemia

Phenothiaziniums as putative photobactericidal agents for red blood cell concentrates.

PubMed

Wainwright, M; Phoenix, D A; Smillie, T E; Wareing, D R

2001-10-01

The antibacterial activities of Methylene Blue and several of its congeners were measured against Yersinia enterocolitica, a gram-negative pathogen known to exhibit significant growth at 4 degrees C and thus constituting a threat to red blood cell concentrates which are stored at this temperature. None of the derivatives was highly active in dark conditions, as expected, but on illumination using a lamp emitting light in the waveband 615-645 nm, considerable bactericidal activity was noted using similar photosensitizer concentrations to those used elsewhere to inactivate blood-borne viruses. Two novel compounds in this area, the phenothiazinium New Methylene Blue N and the phenoxazinium Brilliant Cresyl Blue, exhibited bactericidal activity at lower concentrations than both of the established phenothiaziniums, Methylene Blue and Toluidine Blue O and the recently published blood photovirucidal agent 1,9-Dimethyl Methylene Blue. The photoactivity of these compounds was undiminished in the presence of red blood cells.

Cell wall proteins of Sporothrix schenckii as immunoprotective agents.

PubMed

Alba-Fierro, Carlos A; Pérez-Torres, Armando; López-Romero, Everardo; Cuéllar-Cruz, Mayra; Ruiz-Baca, Estela

2014-01-01

Sporothrix schenckii is the etiological agent of sporotrichosis, an endemic subcutaneous mycosis in Latin America. Cell wall (CW) proteins located on the cell surface are inducers of cellular and humoral immune responses, potential candidates for diagnosis purposes and to generate vaccines to prevent fungal infections. This mini-review emphasizes the potential use of S. schenckii CW proteins as protective and therapeutic immune response inducers against sporotrichosis. A number of pathogenic fungi display CW components that have been characterized as inducers of protective cellular and humoral immune responses against the whole pathogen from which they were originally purified. The isolation and characterization of immunodominant protein components of the CW of S. schenckii have become relevant because of their potential in the development of protective and therapeutic immune responses against sporotrichosis. This manuscript is part of the series of works presented at the "V International Workshop: Molecular genetic approaches to the study of human pathogenic fungi" (Oaxaca, Mexico, 2012). Copyright © 2013 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

Ultrasonic Analysis of Peptide- and Antibody-Targeted Microbubble Contrast Agents for Molecular Imaging of αvβ3-Expressing Cells

PubMed Central

Dayton, Paul A.; Pearson, David; Clark, Jarrod; Simon, Scott; Schumann, Patricia A.; Zutshi, Reena; Matsunaga, Terry O.; Ferrara, Katherine W.

2008-01-01

The goal of targeted ultrasound contrast agents is to significantly and selectively enhance the detection of a targeted vascular site. In this manuscript, three distinct contrast agents targeted to the αvβ3 integrin are examined. The αvβ3 integrin has been shown to be highly expressed on metastatic tumors and endothelial cells during neovascularization, and its expression has been shown to correlate with tumor grade. Specific adhesion of these contrast agents to αvβ3-expressing cell monolayers is demonstrated in vitro, and compared with that of nontargeted agents. Acoustic studies illustrate a backscatter amplitude increase from monolayers exposed to the targeted contrast agents of up to 13-fold (22 dB) relative to enhancement due to control bubbles. A linear dependence between the echo amplitude and bubble concentration was observed for bound agents. The decorrelation of the echo from adherent targeted agents is observed over successive pulses as a function of acoustic pressure and bubble density. Frequency–domain analysis demonstrates that adherent targeted bubbles exhibit high-amplitude narrowband echo components, in contrast to the primarily wideband response from free microbubbles. Results suggest that adherent targeted contrast agents are differentiable from free-floating microbubbles, that targeted contrast agents provide higher sensitivity in the detection of angiogenesis, and that conventional ultrasound imaging techniques such as signal subtraction or decorrelation detection can be used to detect integrin-expressing vasculature with sufficient signal-to-noise. PMID:15296677

Effects of Antiparasite Chemotherapeutic Agents on Immune Functions.

DTIC Science & Technology

1984-05-01

OF ALKYLATING AGENTS AGAINST CELLS PARTICIPATING IN SUPPRESSION OF ANTIBODY RESPONSES* RONALD D. PAUL, ABOUL GHAFFARt and M. MICHAEL SIGEL Department... alkylating agents on the induction and expression of specific suppressor cell activity induced by supraoptimal immunization (SO[) with (4x 109) SRBC was...including 1982b). different subsets of T cells (Cantor & Gershon, A number of alkylating agents which are used as 1979). It is therefore necessary to

A Subset of Latency-Reversing Agents Expose HIV-Infected Resting CD4+ T-Cells to Recognition by Cytotoxic T-Lymphocytes

PubMed Central

Jones, R. Brad; Mueller, Stefanie; O’Connor, Rachel; Rimpel, Katherine; Sloan, Derek D.; Karel, Dan; Wong, Hing C.; Jeng, Emily K.; Thomas, Allison S.; Whitney, James B.; Lim, So-Yon; Kovacs, Colin; Benko, Erika; Karandish, Sara; Huang, Szu-Han; Buzon, Maria J.; Lichterfeld, Mathias; Irrinki, Alivelu; Murry, Jeffrey P.; Tsai, Angela; Yu, Helen; Geleziunas, Romas; Trocha, Alicja; Ostrowski, Mario A.; Irvine, Darrell J.; Walker, Bruce D.

2016-01-01

Resting CD4+ T-cells harboring inducible HIV proviruses are a critical reservoir in antiretroviral therapy (ART)-treated subjects. These cells express little to no viral protein, and thus neither die by viral cytopathic effects, nor are efficiently cleared by immune effectors. Elimination of this reservoir is theoretically possible by combining latency-reversing agents (LRAs) with immune effectors, such as CD8+ T-cells. However, the relative efficacy of different LRAs in sensitizing latently-infected cells for recognition by HIV-specific CD8+ T-cells has not been determined. To address this, we developed an assay that utilizes HIV-specific CD8+ T-cell clones as biosensors for HIV antigen expression. By testing multiple CD8+ T-cell clones against a primary cell model of HIV latency, we identified several single agents that primed latently-infected cells for CD8+ T-cell recognition, including IL-2, IL-15, two IL-15 superagonists (IL-15SA and ALT-803), prostratin, and the TLR-2 ligand Pam3CSK4. In contrast, we did not observe CD8+ T-cell recognition of target cells following treatment with histone deacetylase inhibitors or with hexamethylene bisacetamide (HMBA). In further experiments we demonstrate that a clinically achievable concentration of the IL-15 superagonist ‘ALT-803’, an agent presently in clinical trials for solid and hematological tumors, primes the natural ex vivo reservoir for CD8+ T-cell recognition. Thus, our results establish a novel experimental approach for comparative evaluation of LRAs, and highlight ALT-803 as an LRA with the potential to synergize with CD8+ T-cells in HIV eradication strategies. PMID:27082643

A Subset of Latency-Reversing Agents Expose HIV-Infected Resting CD4+ T-Cells to Recognition by Cytotoxic T-Lymphocytes.

PubMed

Jones, R Brad; Mueller, Stefanie; O'Connor, Rachel; Rimpel, Katherine; Sloan, Derek D; Karel, Dan; Wong, Hing C; Jeng, Emily K; Thomas, Allison S; Whitney, James B; Lim, So-Yon; Kovacs, Colin; Benko, Erika; Karandish, Sara; Huang, Szu-Han; Buzon, Maria J; Lichterfeld, Mathias; Irrinki, Alivelu; Murry, Jeffrey P; Tsai, Angela; Yu, Helen; Geleziunas, Romas; Trocha, Alicja; Ostrowski, Mario A; Irvine, Darrell J; Walker, Bruce D

2016-04-01

Resting CD4+ T-cells harboring inducible HIV proviruses are a critical reservoir in antiretroviral therapy (ART)-treated subjects. These cells express little to no viral protein, and thus neither die by viral cytopathic effects, nor are efficiently cleared by immune effectors. Elimination of this reservoir is theoretically possible by combining latency-reversing agents (LRAs) with immune effectors, such as CD8+ T-cells. However, the relative efficacy of different LRAs in sensitizing latently-infected cells for recognition by HIV-specific CD8+ T-cells has not been determined. To address this, we developed an assay that utilizes HIV-specific CD8+ T-cell clones as biosensors for HIV antigen expression. By testing multiple CD8+ T-cell clones against a primary cell model of HIV latency, we identified several single agents that primed latently-infected cells for CD8+ T-cell recognition, including IL-2, IL-15, two IL-15 superagonists (IL-15SA and ALT-803), prostratin, and the TLR-2 ligand Pam3CSK4. In contrast, we did not observe CD8+ T-cell recognition of target cells following treatment with histone deacetylase inhibitors or with hexamethylene bisacetamide (HMBA). In further experiments we demonstrate that a clinically achievable concentration of the IL-15 superagonist 'ALT-803', an agent presently in clinical trials for solid and hematological tumors, primes the natural ex vivo reservoir for CD8+ T-cell recognition. Thus, our results establish a novel experimental approach for comparative evaluation of LRAs, and highlight ALT-803 as an LRA with the potential to synergize with CD8+ T-cells in HIV eradication strategies.

Quantitative Analyses of Synergistic Responses between Cannabidiol and DNA-Damaging Agents on the Proliferation and Viability of Glioblastoma and Neural Progenitor Cells in Culture.

PubMed

Deng, Liting; Ng, Lindsay; Ozawa, Tatsuya; Stella, Nephi

2017-01-01

Evidence suggests that the nonpsychotropic cannabis-derived compound, cannabidiol (CBD), has antineoplastic activity in multiple types of cancers, including glioblastoma multiforme (GBM). DNA-damaging agents remain the main standard of care treatment available for patients diagnosed with GBM. Here we studied the antiproliferative and cell-killing activity of CBD alone and in combination with DNA-damaging agents (temozolomide, carmustine, or cisplatin) in several human GBM cell lines and in mouse primary GBM cells in cultures. This activity was also studied in mouse neural progenitor cells (NPCs) in culture to assess for potential central nervous system toxicity. We found that CBD induced a dose-dependent reduction of both proliferation and viability of all cells with similar potencies, suggesting no preferential activity for cancer cells. Hill plot analysis indicates an allosteric mechanism of action triggered by CBD in all cells. Cotreatment regimens combining CBD and DNA-damaging agents produced synergistic antiproliferating and cell-killing responses over a limited range of concentrations in all human GBM cell lines and mouse GBM cells as well as in mouse NPCs. Remarkably, antagonistic responses occurred at low concentrations in select human GBM cell lines and in mouse GBM cells. Our study suggests limited synergistic activity when combining CBD and DNA-damaging agents in treating GBM cells, along with little to no therapeutic window when considering NPCs. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

Interactome Analysis of Microtubule-targeting Agents Reveals Cytotoxicity Bases in Normal Cells.

PubMed

Gutiérrez-Escobar, Andrés Julián; Méndez-Callejas, Gina

2017-12-01

Cancer causes millions of deaths annually and microtubule-targeting agents (MTAs) are the most commonly-used anti-cancer drugs. However, the high toxicity of MTAs on normal cells raises great concern. Due to the non-selectivity of MTA targets, we analyzed the interaction network in a non-cancerous human cell. Subnetworks of fourteen MTAs were reconstructed and the merged network was compared against a randomized network to evaluate the functional richness. We found that 71.4% of the MTA interactome nodes are shared, which affects cellular processes such as apoptosis, cell differentiation, cell cycle control, stress response, and regulation of energy metabolism. Additionally, possible secondary targets were identified as client proteins of interphase microtubules. MTAs affect apoptosis signaling pathways by interacting with client proteins of interphase microtubules, suggesting that their primary targets are non-tumor cells. The paclitaxel and doxorubicin networks share essential topological axes, suggesting synergistic effects. This may explain the exacerbated toxicity observed when paclitaxel and doxorubicin are used in combination for cancer treatment. Copyright © 2017 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

Improved Lubricating Performance by Combining Oil-Soluble Hairy Silica Nanoparticles and an Ionic Liquid as an Additive for a Synthetic Base Oil

DOE PAGES

Seymour, Bryan T.; Fu, Wenxin; Wright, Roger A. E.; ...

2018-04-05

This article reports on improved lubricating performance by combining oil-soluble poly(lauryl methacrylate) brush-grafted silica nanoparticles (hairy NPs or HNP) and an oil-miscible phosphonium-phosphate ionic liquid (IL) as a friction-reducing additive for a polyalphaolefin (PAO) oil. The HNP was synthesized by surface-initiated reversible addition–fragmentation chain transfer polymerization. At a total concentration of 2% and sufficiently high individual concentrations for HNP and IL in PAO, high-contact stress, ball-on-flat reciprocating tribological tests showed that the friction decreased by up to 23% compared with 2% HNP alone in PAO and by up to 35% compared to the PAO mixed with 2% IL. Scanning electronmore » microscopy, energy-dispersive X-ray spectroscopy, and X-ray photoelectron spectroscopy (XPS) analysis revealed that the tribofilm formed from the PAO containing 1% HNP + 1% IL was enriched with both Si and P, indicating that both hairy NPs and IL were involved in the tribochemical reactions. In addition, the O 1s and Si 2p peaks in the core-level XPS spectra exhibited significant shifts for the mixture of 1% HNP + 1% IL compared to those for 2% HNP, suggesting the possible formation of new covalent bonds. These results indicated that HNP and IL reacted with each other and also with the metal substrate during the rubbing process, which likely strengthened the tribofilm and its bonding with the substrate and thus further improved the lubrication.« less

Improved Lubricating Performance by Combining Oil-Soluble Hairy Silica Nanoparticles and an Ionic Liquid as an Additive for a Synthetic Base Oil

DOE Office of Scientific and Technical Information (OSTI.GOV)

Seymour, Bryan T.; Fu, Wenxin; Wright, Roger A. E.

This article reports on improved lubricating performance by combining oil-soluble poly(lauryl methacrylate) brush-grafted silica nanoparticles (hairy NPs or HNP) and an oil-miscible phosphonium-phosphate ionic liquid (IL) as a friction-reducing additive for a polyalphaolefin (PAO) oil. The HNP was synthesized by surface-initiated reversible addition–fragmentation chain transfer polymerization. At a total concentration of 2% and sufficiently high individual concentrations for HNP and IL in PAO, high-contact stress, ball-on-flat reciprocating tribological tests showed that the friction decreased by up to 23% compared with 2% HNP alone in PAO and by up to 35% compared to the PAO mixed with 2% IL. Scanning electronmore » microscopy, energy-dispersive X-ray spectroscopy, and X-ray photoelectron spectroscopy (XPS) analysis revealed that the tribofilm formed from the PAO containing 1% HNP + 1% IL was enriched with both Si and P, indicating that both hairy NPs and IL were involved in the tribochemical reactions. In addition, the O 1s and Si 2p peaks in the core-level XPS spectra exhibited significant shifts for the mixture of 1% HNP + 1% IL compared to those for 2% HNP, suggesting the possible formation of new covalent bonds. These results indicated that HNP and IL reacted with each other and also with the metal substrate during the rubbing process, which likely strengthened the tribofilm and its bonding with the substrate and thus further improved the lubrication.« less

Characterisation of a Cell Culture System for Investigating Nerve Agent Neurotoxicology. Part 1

DTIC Science & Technology

2012-03-01

eds. CRC Press), pp. 1-24. Sawyer,T.W., Weiss,M.T., and Unger,R.J. (1992). Anticholinesterase activity of organophosphate nerve agents in neuronal...were confirmed. The presence of muscarinic receptors and acetylcholinesterase activity was determined. Importantly, differential acetylcholinesterase... activity assays that will provide the basis for an ongoing research programme. The neuroblastoma cell lines chosen can potentially be used as a

Nanoparticles increase the efficacy of cancer chemopreventive agents in cells exposed to cigarette smoke condensate.

PubMed

Pulliero, Alessandra; Wu, Yun; Fenoglio, Daniela; Parodi, Alessia; Romani, Massimo; Soares, Christiane P; Filaci, Gilberto; Lee, James L; Sinkam, Patrick N; Izzotti, Alberto

2015-03-01

Lung cancer is a leading cause of death in developed countries. Although smoking cessation is a primary strategy for preventing lung cancer, former smokers remain at high risk of cancer. Accordingly, there is a need to increase the efficacy of lung cancer prevention. Poor bioavailability is the main factor limiting the efficacy of chemopreventive agents. The aim of this study was to increase the efficacy of cancer chemopreventive agents by using lipid nanoparticles (NPs) as a carrier. This study evaluated the ability of lipid NPs to modify the pharmacodynamics of chemopreventive agents including N-acetyl-L-cysteine, phenethyl isothiocyanate and resveratrol (RES). The chemopreventive efficacy of these drugs was determined by evaluating their abilities to counteract cytotoxic damage (DNA fragmentation) induced by cigarette smoke condensate (CSC) and to activate protective apoptosis (annexin-V cytofluorimetric staining) in bronchial epithelial cells both in vitro and in ex vivo experiment in mice. NPs decreased the toxicity of RES and increased its ability to counteract CSC cytotoxicity. NPs significantly increased the ability of phenethyl isothiocyanate to attenuate CSC-induced DNA fragmentation at the highest tested dose. In contrast, this potentiating effect was observed at all tested doses of RES, NPs dramatically increasing RES-induced apoptosis in CSC-treated cells. These results provide evidence that NPs are highly effective at increasing the efficacy of lipophilic drugs (RES) but are not effective towards hydrophilic agents (N-acetyl-L-cysteine), which already possess remarkable bioavailability. Intermediate effects were observed for phenethyl isothiocyanate. These findings are relevant to the identification of cancer chemopreventive agents that would benefit from lipid NP delivery. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Use of trimetasphere metallofullerene MRI contrast agent for the non-invasive longitudinal tracking of stem cells in the lung

PubMed Central

Murphy, Sean V.; Hale, Austin; Reid, Tanya; Olson, John; Kidiyoor, Amritha; Tan, Josh; Zhou, Zhiguo; Jackson, John; Atala, Anthony

2016-01-01

Magnetic Resonance Imaging (MRI) is a commonly used, non-invasive imaging technique that provides visualization of soft tissues with high spatial resolution. In both a research and clinical setting, the major challenge has been identifying a non-invasive and safe method for longitudinal tracking of delivered cells in vivo. The labeling and tracking of contrast agent labeled cells using MRI has the potential to fulfill this need. Contrast agents are often used to enhance the image contrast between the tissue of interest and surrounding tissues with MRI. The most commonly used MRI contrast agents contain Gd(III) ions. However, Gd(III) ions are highly toxic in their ionic form, as they tend to accumulate in the liver, spleen, kidney and bones and block calcium channels. Endohedral metallofullerenes such as trimetallic nitride endohedral metallofullerenes (Trimetasphere®) are one unique class of fullerene molecules where a Gd3N cluster is encapsulated inside a C80 carbon cage referred to as Gd3N@C80. These endohedral metallofullerenes have several advantages over small chelated Gd(III) complexes such as increased stability of the Gd(III) ion, minimal toxic effects, high solubility in water and high proton relativity. In this study, we describe the evaluation of gadolinium-based Trimetasphere® positive contrast agent for the in vitro labeling and in vivo tracking of human amniotic fluid-derived stem cells within lung tissue. In addition, we conducted a ‘proof-of-concept’ experiment demonstrating that this methodology can be used to track the homing of stem cells to injured lung tissue and provide longitudinal analysis of cell localization over an extended time course. PMID:26546729

Use of trimetasphere metallofullerene MRI contrast agent for the non-invasive longitudinal tracking of stem cells in the lung.

PubMed

Murphy, Sean V; Hale, Austin; Reid, Tanya; Olson, John; Kidiyoor, Amritha; Tan, Josh; Zhou, Zhiguo; Jackson, John; Atala, Anthony

2016-04-15

Magnetic Resonance Imaging (MRI) is a commonly used, non-invasive imaging technique that provides visualization of soft tissues with high spatial resolution. In both a research and clinical setting, the major challenge has been identifying a non-invasive and safe method for longitudinal tracking of delivered cells in vivo. The labeling and tracking of contrast agent labeled cells using MRI has the potential to fulfill this need. Contrast agents are often used to enhance the image contrast between the tissue of interest and surrounding tissues with MRI. The most commonly used MRI contrast agents contain Gd(III) ions. However, Gd(III) ions are highly toxic in their ionic form, as they tend to accumulate in the liver, spleen, kidney and bones and block calcium channels. Endohedral metallofullerenes such as trimetallic nitride endohedral metallofullerenes (Trimetasphere®) are one unique class of fullerene molecules where a Gd3N cluster is encapsulated inside a C80 carbon cage referred to as Gd3N@C80. These endohedral metallofullerenes have several advantages over small chelated Gd(III) complexes such as increased stability of the Gd(III) ion, minimal toxic effects, high solubility in water and high proton relativity. In this study, we describe the evaluation of gadolinium-based Trimetasphere® positive contrast agent for the ​in vitro labeling and in vivo tracking of human amniotic fluid-derived stem cells within lung tissue. In addition, we conducted a 'proof-of-concept' experiment demonstrating that this methodology can be used to track the homing of stem cells to injured lung tissue and provide longitudinal analysis of cell localization over an extended time course. Copyright © 2015 Elsevier Inc. All rights reserved.

Glucose-installed, SPIO-loaded PEG- b-PCL micelles as MR contrast agents to target prostate cancer cells

NASA Astrophysics Data System (ADS)

Theerasilp, Man; Sunintaboon, Panya; Sungkarat, Witaya; Nasongkla, Norased

2017-11-01

Polymeric micelles of poly(ethylene glycol)- block-poly(ɛ-caprolactone) bearing glucose analog encapsulated with superparamagnetic iron oxide nanoparticles (Glu-SPIO micelles) were synthesized as an MRI contrast agent to target cancer cells based on high-glucose metabolism. Compared to SPIO micelles (non-targeting SPIO micelles), Glu-SPIO micelles demonstrated higher toxicity to human prostate cancer cell lines (PC-3) at high concentration. Atomic absorption spectroscopy was used to determine the amount of iron in cells. It was found that the iron in cancer cells treated by Glu-SPIO micelles were 27-fold higher than cancer cells treated by SPIO micelles at the iron concentration of 25 ppm and fivefold at the iron concentration of 100 ppm. To implement Glu-SPIO micelles as a MR contrast agent, the 3-T clinical MRI was applied to determine transverse relaxivities ( r 2*) and relaxation rate (1/ T 2*) values. In vitro MRI showed different MRI signal from cancer cells after cellular uptake of SPIO micelles and Glu-SPIO micelles. Glu-SPIO micelles was highly sensitive with the r 2* in agarose gel at 155 mM-1 s-1. Moreover, the higher 1/ T 2* value was found for cancer cells treated with Glu-SPIO micelles. These results supported that glucose ligand increased the cellular uptake of micelles by PC-3 cells with over-expressing glucose transporter on the cell membrane. Thus, glucose can be used as a small molecule ligand for targeting prostate cancer cells overexpressing glucose transporter.

Unipotent, Atoh1+ progenitors maintain the Merkel cell population in embryonic and adult mice

PubMed Central

Wright, Margaret C.; Reed-Geaghan, Erin G.; Bolock, Alexa M.; Fujiyama, Tomoyuki; Hoshino, Mikio

2015-01-01

Resident progenitor cells in mammalian skin generate new cells as a part of tissue homeostasis. We sought to identify the progenitors of Merkel cells, a unique skin cell type that plays critical roles in mechanosensation. We found that some Atoh1-expressing cells in the hairy skin and whisker follicles are mitotically active at embryonic and postnatal ages. Genetic fate-mapping revealed that these Atoh1-expressing cells give rise solely to Merkel cells. Furthermore, selective ablation of Atoh1+ skin cells in adult mice led to a permanent reduction in Merkel cell numbers, demonstrating that other stem cell populations are incapable of producing Merkel cells. These data identify a novel, unipotent progenitor population in the skin that gives rise to Merkel cells both during development and adulthood. PMID:25624394

Selected Alkylating Agents Can Overcome Drug Tolerance of G0-like Tumor Cells and Eradicate BRCA1-Deficient Mammary Tumors in Mice.

PubMed

Pajic, Marina; Blatter, Sohvi; Guyader, Charlotte; Gonggrijp, Maaike; Kersbergen, Ariena; Küçükosmanoğlu, Aslι; Sol, Wendy; Drost, Rinske; Jonkers, Jos; Borst, Piet; Rottenberg, Sven

2017-11-15

Purpose: We aimed to characterize and target drug-tolerant BRCA1-deficient tumor cells that cause residual disease and subsequent tumor relapse. Experimental Design: We studied responses to various mono- and bifunctional alkylating agents in a genetically engineered mouse model for BRCA1/p53 -mutant breast cancer. Because of the large intragenic deletion of the Brca1 gene, no restoration of BRCA1 function is possible, and therefore, no BRCA1-dependent acquired resistance occurs. To characterize the cell-cycle stage from which Brca1 -/- ;p53 -/- mammary tumors arise after cisplatin treatment, we introduced the fluorescent ubiquitination-based cell-cycle indicator (FUCCI) construct into the tumor cells. Results: Despite repeated sensitivity to the MTD of platinum drugs, the Brca1 -mutated mammary tumors are not eradicated, not even by a frequent dosing schedule. We show that relapse comes from single-nucleated cells delaying entry into the S-phase. Such slowly cycling cells, which are present within the drug-naïve tumors, are enriched in tumor remnants. Using the FUCCI construct, we identified nonfluorescent G 0 -like cells as the population most tolerant to platinum drugs. Intriguingly, these cells are more sensitive to the DNA-crosslinking agent nimustine, resulting in an increased number of multinucleated cells that lack clonogenicity. This is consistent with our in vivo finding that the nimustine MTD, among several alkylating agents, is the most effective in eradicating Brca1 -mutated mouse mammary tumors. Conclusions: Our data show that targeting G 0 -like cells is crucial for the eradication of BRCA1/p53-deficient tumor cells. This can be achieved with selected alkylating agents such as nimustine. Clin Cancer Res; 23(22); 7020-33. ©2017 AACR . ©2017 American Association for Cancer Research.

Phylogeography of screaming hairy armadillo Chaetophractus vellerosus: Successive disjunctions and extinctions due to cyclical climatic changes in southern South America

PubMed Central

Ferreiro, Alejandro M.; Chiappero, Marina B.; Sánchez, Julieta; Gabrielli, Magalí; Lizarralde, Marta S.

2018-01-01

Little is known about phylogeography of armadillo species native to southern South America. In this study we describe the phylogeography of the screaming hairy armadillo Chaetophractus vellerosus, discuss previous hypothesis about the origin of its disjunct distribution and propose an alternative one, based on novel information on genetic variability. Variation of partial sequences of mitochondrial DNA Control Region (CR) from 73 individuals from 23 localities were analyzed to carry out a phylogeographic analysis using neutrality tests, mismatch distribution, median-joining (MJ) network and paleontological records. We found 17 polymorphic sites resulting in 15 haplotypes. Two new geographic records that expand known distribution of the species are presented; one of them links the distributions of recently synonimized species C. nationi and C. vellerosus. Screaming hairy armadillo phylogeographic pattern can be addressed as category V of Avise: common widespread linages plus closely related lineages confined to one or a few nearby locales each. The older linages are distributed in the north-central area of the species distribution range in Argentina (i.e. ancestral area of distribution). C. vellerosus seems to be a low vagility species that expanded, and probably is expanding, its distribution range while presents signs of genetic structuring in central areas. To explain the disjunct distribution, a hypothesis of extinction of the species in intermediate areas due to quaternary climatic shift to more humid conditions was proposed. We offer an alternative explanation: long distance colonization, based on null genetic variability, paleontological record and evidence of alternance of cold/arid and temperate/humid climatic periods during the last million years in southern South America. PMID:29324793

Highly versatile SPION encapsulated PLGA nanoparticles as photothermal ablators of cancer cells and as multimodal imaging agents.

PubMed

Sivakumar, Balasubramanian; Aswathy, Ravindran Girija; Romero-Aburto, Rebeca; Mitcham, Trevor; Mitchel, Keith A; Nagaoka, Yutaka; Bouchard, Richard R; Ajayan, Pulickel M; Maekawa, Toru; Sakthikumar, Dasappan Nair

2017-02-28

We have designed versatile polymeric nanoparticles with cancer cell specific targeting capabilities via aptamer conjugation after the successful encapsulation of curcumin and superparamagnetic iron oxide nanoparticles (SPIONs) inside a PLGA nanocapsule. These targeted nanocomposites were selectively taken up by tumor cells, under in vitro conditions, demonstrating the effectiveness of the aptamer targeting mechanism. Moreover, the nanocomposite potentially functioned as efficient multiprobes for optical, magnetic resonance imaging (MRI) and photoacoustic imaging contrast agents in the field of cancer diagnostics. The hyperthermic ability of these nanocomposites was mediated by SPIONs upon NIR-laser irradiation. In vitro cytotoxicity was shown by curcumin-loaded nanoparticles as well as the photothermal ablation of cancer cells mediated by the drug-encapsulated nanocomposite demonstrated the potential therapeutic effect of the nanocomposite. In short, we portray the aptamer-conjugated nanocomposite as a multimodal material capable of serving as a contrast agent for MR, photoacoustic and optical imaging. Furthermore, the nanocomposite functions as a targetable drug nanocarrier and a NIR-laser inducible hyperthermic material that is capable of ablating PANC-1 and MIA PaCa-2 cancer cell lines.

Antibody-Directed Cytotoxic Agents: Use of Monoclonal Antibody to Direct the Action of Toxin A Chains to Colorectal Carcinoma Cells

NASA Astrophysics Data System (ADS)

Gilliland, D. Gary; Steplewski, Zenon; Collier, R. John; Mitchell, Kenneth F.; Chang, Tong H.; Koprowski, Hilary

1980-08-01

We have constructed cell-specific cytotoxic agents by covalently coupling the A chain from diphtheria toxin or ricin toxin to monoclonal antibody directed against a colorectal carcinoma tumor-associated antigen. Antibody 1083-17-1A was modified by attachment of 3-(2-pyridyldithio)propionyl or cystaminyl groups and then treated with reduced A chain to give disulfide-linked conjugates that retained the original binding specificity of the antibody moiety. The conjugates showed cytotoxic activity for colorectal carcinoma cells in culture, but were not toxic in the same concentration range for a variety of cell lines that lacked the antigen. Under defined conditions virtually 100% of antigen-bearing cultured cells were killed, whereas cells that lacked the antigen were not affected. Conjugates containing toxin A chains coupled to monoclonal antibodies may be useful in studying functions of various cell surface components and, possibly, as tumor-specific therapeutic agents.

The enhanced inhibitory effect of different antitumor agents in self-microemulsifying drug delivery systems on human cervical cancer HeLa cells.

PubMed

Ujhelyi, Zoltán; Kalantari, Azin; Vecsernyés, Miklós; Róka, Eszter; Fenyvesi, Ferenc; Póka, Róbert; Kozma, Bence; Bácskay, Ildikó

2015-07-21

The aim of this study was to develop topical self-microemulsifying drug delivery systems (SMEDDS) containing antitumor agents (bleomycin, cisplatin and ifosfamide) and to investigate their inhibitory potential in SMEDDS on human cervical cancer HeLa cells. The physicochemical properties of cytostatic drug loaded SMEDDS were characterized. The cytotoxicity of main components of SMEDDS was also investigated. Their IC50 values were determined. HeLa cells were treated by different concentrations of cisplatin, bleomycin and ifosfamide alone and in various SMEDDS. The inhibitory effect on cell growth was analyzed by MTT cell viability assay. Inflammation is a driving force that accelerates cancer development. The inhibitory effect of these antitumor agents has also been tested on HeLa cells in the presence of inflammatory mediators (IL-1-β, TNF-α) as an in vitro model of inflamed human cervix. Significant differences in the cytotoxicity of cytostatic drugs alone and in SMEDDS have been found in a concentration-dependent manner. The self-micro emulsifying system may potentiate the effectiveness of bleomycin, cisplatin and ifosfamide topically. The effect of SMEDDS containing antitumor agents was decreased significantly in the presence of inflammatory mediators. According to our experiments, the optimal SMEDDS formulation is 1:1:2:6:2 ratios of Isopropyl myristate, Capryol 90, Kolliphor RH 40, Cremophor RH40, Transcutol HP and Labrasol. It can be concluded that SMEDDS may increase the inhibitory effect of bleomycin, ifosfamide and cisplatin on human cervical cancer HeLa cells. Inflammation on HeLa cells hinders the effectiveness of SMEDDS containing antitumor agents. Our results might ensure useful data for development of optimal antitumor formulations.

Influence of molecular weight on the phase behavior and structure formation of branched side-chain hairy-rod polyfluorene in bulk phase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Knaapila, M.; Lyons, B.P.; Foreman, J.P.

We report on an experimental study of the self-organization and phase behavior of hairy-rod {pi}-conjugated branched side-chain polyfluorene, poly[9,9-bis(2-ethylhexyl)-fluorene-2,7-diyl] - i.e., poly[2,7-(9,9-bis(2-ethylhexyl)fluorene] (PF2/6) - as a function of molecular weight (M{sub n}). The results have been compared to those of phenomenological theory. Samples for which M{sub n}=3-147 kg/mol were used. First, the stiffness of PF2/6, the assumption of the theory, has been probed by small-angle neutron scattering in solution. Thermogravimetry has been used to show that PF2/6 is thermally stable over the conditions studied. Second, the existence of nematic and hexagonal phases has been phenomenologically identified for lower and highermore » M{sub n} (LMW, M{sub n}M{sub n}{sup *}) regimes, respectively, based on free-energy argument of nematic and hexagonal hairy rods and found to correspond to the experimental x-ray diffraction (XRD) results for PF2/6. By using the lattice parameters of PF2/6 as an experimental input, the nematic-hexagonal transition has been predicted in the vicinity of glassification temperature (T{sub g}) of PF2/6. Then, by taking the orientation parts of the free energies into account the nematic-hexagonal transition has been calculated as a function of temperature and M{sub n} and a phase diagram has been formed. Below T{sub g} of 80 deg. C only (frozen) nematic phase is observed for M{sub n}M{sub n}*. The nematic-hexagonal transition upon heating is observed for the HMW regime depending weakly on M{sub n}, being at 140-165 deg. C for M{sub n}>M{sub n}*. Third, the phase behavior and structure formation as a function of M{sub n} have been probed using powder and fiber XRD and differential scanning calorimetry and reasonable semiquantitative agreement with theory has been found for M{sub n}{>=}3 kg/mol. Fourth, structural characteristics

Method for enhancing cell penetration of Gd3+-based MRI contrast agents by conjugation with hydrophobic fluorescent dyes.

PubMed

Yamane, Takehiro; Hanaoka, Kenjiro; Muramatsu, Yasuaki; Tamura, Keita; Adachi, Yusuke; Miyashita, Yasushi; Hirata, Yasunobu; Nagano, Tetsuo

2011-11-16

Gadolinium ion (Gd(3+)) complexes are commonly used as magnetic resonance imaging (MRI) contrast agents to enhance signals in T(1)-weighted MR images. Recently, several methods to achieve cell-permeation of Gd(3+) complexes have been reported, but more general and efficient methodology is needed. In this report, we describe a novel method to achieve cell permeation of Gd(3+) complexes by using hydrophobic fluorescent dyes as a cell-permeability-enhancing unit. We synthesized Gd(3+) complexes conjugated with boron dipyrromethene (BDP-Gd) and Cy7 dye (Cy7-Gd), and showed that these conjugates can be introduced efficiently into cells. To examine the relationship between cell permeability and dye structure, we further synthesized a series of Cy7-Gd derivatives. On the basis of MR imaging, flow cytometry, and ICP-MS analysis of cells loaded with Cy7-Gd derivatives, highly hydrophobic and nonanionic dyes were effective for enhancing cell permeation of Gd(3+) complexes. Furthermore, the behavior of these Cy7-Gd derivatives was examined in mice. Thus, conjugation of hydrophobic fluorescent dyes appears to be an effective approach to improve the cell permeability of Gd(3+) complexes, and should be applicable for further development of Gd(3+)-based MRI contrast agents.

Skin stem cell hypotheses and long term clone survival--explored using agent-based modelling.

PubMed

Li, X; Upadhyay, A K; Bullock, A J; Dicolandrea, T; Xu, J; Binder, R L; Robinson, M K; Finlay, D R; Mills, K J; Bascom, C C; Kelling, C K; Isfort, R J; Haycock, J W; MacNeil, S; Smallwood, R H

2013-01-01

Epithelial renewal in skin is achieved by the constant turnover and differentiation of keratinocytes. Three popular hypotheses have been proposed to explain basal keratinocyte regeneration and epidermal homeostasis: 1) asymmetric division (stem-transit amplifying cell); 2) populational asymmetry (progenitor cell with stochastic fate); and 3) populational asymmetry with stem cells. In this study, we investigated lineage dynamics using these hypotheses with a 3D agent-based model of the epidermis. The model simulated the growth and maintenance of the epidermis over three years. The offspring of each proliferative cell was traced. While all lineages were preserved in asymmetric division, the vast majority were lost when assuming populational asymmetry. The third hypothesis provided the most reliable mechanism for self-renewal by preserving genetic heterogeneity in quiescent stem cells, and also inherent mechanisms for skin ageing and the accumulation of genetic mutation.

Cellular effects of the microtubule-targeting agent peloruside A in hypoxia-conditioned colorectal carcinoma cells.

PubMed

Řehulka, Jiří; Annadurai, Narendran; Frydrych, Ivo; Znojek, Pawel; Džubák, Petr; Northcote, Peter; Miller, John H; Hajdúch, Marián; Das, Viswanath

2017-07-01

Hypoxia is a prominent feature of solid tumors, dramatically remodeling microtubule structures and cellular pathways and contributing to paclitaxel resistance. Peloruside A (PLA), a microtubule-targeting agent, has shown promising anti-tumor effects in preclinical studies. Although it has a similar mode of action to paclitaxel, it binds to a distinct site on β-tubulin that differs from the classical taxane site. In this study, we examined the unexplored effects of PLA in hypoxia-conditioned colorectal HCT116 cancer cells. Cytotoxicity of PLA was determined by cell proliferation assay. The effects of a pre-exposure to hypoxia on PLA-induced cell cycle alterations and apoptosis were examined by flow cytometry, time-lapse imaging, and western blot analysis of selected markers. The hypoxia effect on stabilization of microtubules by PLA was monitored by an intracellular tubulin polymerization assay. Our findings show that the cytotoxicity of PLA is not altered in hypoxia-conditioned cells compared to paclitaxel and vincristine. Furthermore, hypoxia does not alter PLA-induced microtubule stabilization nor the multinucleation of cells. PLA causes cyclin B1 and G2/M accumulation followed by apoptosis. The cellular and molecular effects of PLA have been determined in normoxic conditions, but there are no reports of PLA effects in hypoxic cells. Our findings reveal that hypoxia preconditioning does not alter the sensitivity of HCT116 to PLA. These data report on the cellular and molecular effects of PLA in hypoxia-conditioned cells for the first time, and will encourage further exploration of PLA as a promising anti-tumor agent. Copyright © 2017 Elsevier B.V. All rights reserved.

Intracellular haemolytic agents of Heterocapsa circularisquama exhibit toxic effects on H. circularisquama cells themselves and suppress both cell-mediated haemolytic activity and toxicity to rotifers (Brachionus plicatilis).

PubMed

Nishiguchi, Tomoki; Cho, Kichul; Yasutomi, Masumi; Ueno, Mikinori; Yamaguchi, Kenichi; Basti, Leila; Yamasaki, Yasuhiro; Takeshita, Satoshi; Kim, Daekyung; Oda, Tatsuya

2016-10-01

A harmful dinoflagellate, Heterocapsa circularisquama, is highly toxic to shellfish and the zooplankton rotifer Brachionus plicatilis. A previous study found that H. circularisquama has both light-dependent and -independent haemolytic agents, which might be responsible for its toxicity. Detailed analysis of the haemolytic activity of H. circularisquama suggested that light-independent haemolytic activity was mediated mainly through intact cells, whereas light-dependent haemolytic activity was mediated by intracellular agents which can be discharged from ruptured cells. Because H. circularisquama showed similar toxicity to rotifers regardless of the light conditions, and because ultrasonic ruptured H. circularisquama cells showed no significant toxicity to rotifers, it was suggested that live cell-mediated light-independent haemolytic activity is a major factor responsible for the observed toxicity to rotifers. Interestingly, the ultrasonic-ruptured cells of H. circularisquama suppressed their own lethal effect on the rotifers. Analysis of samples of the cell contents (supernatant) and cell fragments (precipitate) prepared from the ruptured H. circularisquama cells indicated that the cell contents contain inhibitors for the light-independent cell-mediated haemolytic activity, toxins affecting H. circularisquama cells themselves, as well as light-dependent haemolytic agents. Ethanol extract prepared from H. circularisquama, which is supposed to contain a porphyrin derivative that displays photosensitising haemolytic activity, showed potent toxicity to Chattonella marina, Chattonella antiqua, and Karenia mikimotoi, as well as to H. circularisquama at the concentration range at which no significant toxicity to rotifers was observed. Analysis on a column of Sephadex LH-20 revealed that light-dependent haemolytic activity and inhibitory activity on cell-mediated light-independent haemolytic activity existed in two separate fractions (f-2 and f-3), suggesting that both

Alisertib in Combination With Vorinostat in Treating Patients With Relapsed or Recurrent Hodgkin Lymphoma, B-Cell Non-Hodgkin Lymphoma, or Peripheral T-Cell Lymphoma

ClinicalTrials.gov

2018-04-10

Adult B Acute Lymphoblastic Leukemia; Adult T Acute Lymphoblastic Leukemia; Anaplastic Large Cell Lymphoma; Angioimmunoblastic T-Cell Lymphoma; Chronic Lymphocytic Leukemia; Extranodal Marginal Zone Lymphoma of Mucosa-Associated Lymphoid Tissue; Hepatosplenic T-Cell Lymphoma; Intraocular Lymphoma; Lymphomatous Involvement of Non-Cutaneous Extranodal Site; Mature T-Cell and NK-Cell Non-Hodgkin Lymphoma; Nodal Marginal Zone Lymphoma; Primary Cutaneous B-Cell Non-Hodgkin Lymphoma; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Adult Burkitt Lymphoma; Recurrent Adult Grade III Lymphomatoid Granulomatosis; Recurrent Adult Hodgkin Lymphoma; Recurrent Adult Immunoblastic Lymphoma; Recurrent Adult Lymphoblastic Lymphoma; Recurrent Adult T-Cell Leukemia/Lymphoma; Recurrent Grade 1 Follicular Lymphoma; Recurrent Grade 2 Follicular Lymphoma; Recurrent Grade 3 Follicular Lymphoma; Recurrent Mantle Cell Lymphoma; Recurrent Marginal Zone Lymphoma; Recurrent Mycosis Fungoides and Sezary Syndrome; Recurrent Non-Hodgkin Lymphoma; Recurrent Primary Cutaneous T-Cell Non-Hodgkin Lymphoma; Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; Refractory Hairy Cell Leukemia; Small Intestinal Lymphoma; Splenic Marginal Zone Lymphoma; T-Cell Large Granular Lymphocyte Leukemia; Testicular Lymphoma; Waldenstrom Macroglobulinemia

(+)-Grandifloracin, an antiausterity agent, induces autophagic PANC-1 pancreatic cancer cell death.

PubMed

Ueda, Jun-ya; Athikomkulchai, Sirivan; Miyatake, Ryuta; Saiki, Ikuo; Esumi, Hiroyasu; Awale, Suresh

2014-01-01

Human pancreatic tumors are known to be highly resistant to nutrient starvation, and this prolongs their survival in the hypovascular (austere) tumor microenvironment. Agents that retard this tolerance to nutrient starvation represent a novel antiausterity strategy in anticancer drug discovery. (+)-Grandifloracin (GF), isolated from Uvaria dac, has shown preferential toxicity to PANC-1 human pancreatic cancer cells under nutrient starvation, with a PC50 value of 14.5 μM. However, the underlying mechanism is not clear. In this study, GF was found to preferentially induce PANC-1 cell death in a nutrient-deprived medium via hyperactivation of autophagy, as evidenced by a dramatic upregulation of microtubule-associated protein 1 light chain 3. No change was observed in expression of the caspase-3 and Bcl-2 apoptosis marker proteins. GF was also found to strongly inhibit the activation of Akt, a key regulator of cancer cell survival and proliferation. Because pancreatic tumors are highly resistant to current therapies that induce apoptosis, the alternative cell death mechanism exhibited by GF provides a novel therapeutic insight into antiausterity drug candidates.

Langerhans Cells and Their Role in Oral Mucosal Diseases

PubMed Central

Upadhyay, Juhi; Upadhyay, Ram B; Agrawal, Pankaj; Jaitley, Shweta; Shekhar, Rhitu

2013-01-01

Dendritic cells are arguably the most potent antigen-presenting cells and may be the only cells capable of initiating the adaptive immune response. The epithelial residents of dendritic cells are Langerhans cells, which serve as the “sentinels” of the mucosa, altering the immune system not only to pathogen entry but also of tolerance to self antigen and commensal microbes. Oral mucosal Langerhans cells are capable of engaging and internalizing a wide variety of pathogens and have been found responsive to nickel in patients with nickel allergies, oral Candida species, oral lichen planus, lichenoid drug eruptions, graft versus host diseases, periodontal diseases median rhomboid glossitis, human immunodeficiency virus infection, hairy leukoplakia of the tongue, and oral squamous cell carcinoma. Review focuses on the role of antigen-presenting cells in particular Langerhans cells to better understand the mechanisms underlying immune responses. In this review, comprehensive detail about mucosal diseases has been compiled using the PubMed database and through textbooks. PMID:24251267

Quantifying cancer cell receptors with paired-agent fluorescent imaging: a novel method to account for tissue optical property effects

NASA Astrophysics Data System (ADS)

Sadeghipour, Negar; Davis, Scott C.; Tichauer, Kenneth M.

2018-02-01

Dynamic fluorescence imaging approaches can be used to estimate the concentration of cell surface receptors in vivo. Kinetic models are used to generate the final estimation by taking the targeted imaging agent concentration as a function of time. However, tissue absorption and scattering properties cause the final readout signal to be on a different scale than the real fluorescent agent concentration. In paired-agent imaging approaches, simultaneous injection of a suitable control imaging agent with a targeted one can account for non-specific uptake and retention of the targeted agent. Additionally, the signal from the control agent can be a normalizing factor to correct for tissue optical property differences. In this study, the kinetic model used for paired-agent imaging analysis (i.e., simplified reference tissue model) is modified and tested in simulation and experimental data in a way that accounts for the scaling correction within the kinetic model fit to the data to ultimately extract an estimate of the targeted biomarker concentration.

Identification of cancer specific ligands from one-bead one compound combinatorial libraries to develop theranostics agents against oral squamous cell carcinoma

NASA Astrophysics Data System (ADS)

Yang, Frances Fan

Background: Oral squamous cell carcinoma (OSCC) is one of the most prevalent disease worldwide. One-bead one-compound (OBOC) combinatorial technology is a powerful method to identify peptidomimetic ligands against a variety of receptors on cell surfaces. We therefore hypothesized that cancer specific ligands against OSCC might be identified and can be conjugated to optical dyes or nanocarriers to develop theranostic agents against OSCC. Material and methods: Different OSCC cell lines were incubated with OBOC libraries and beads with cell binding were sorted and then screened with normal human cells to identify peptide-beads binding to different OSCC cell lines but not binding to normal human cells. The molecular probes of OSCC were developed by biotinylating the carboxyl end of the ligands. OSCC theranostic agents were developed by decorating LLY13 with NPs and evaluated by using orthotopic bioluminescent oral cancer model. Results: Six OSCC specific ligands were discovered. Initial peptide-histochemistry study indicated that LLY12 and LLY13 were able to specifically detect OSCC cells grown on chamber slides at the concentration of 1 muM. In addition, LLY13 was found to penetrate into the OSCC cells and accumulate in the cytoplasm, and nucleus. After screened with a panel of integrin antibodies, only anti-alpha3 antibody was able to block most of OSCC cells binding to the LLY13 beads. OSCC theranostic agents developed using targeting LLY13 micelles (25+/- 4nm in diameter) were more efficient in binding to HSC-3 cancer cells compared to non-targeting micelles. Ex vivo images demonstrated that xenografts from the mice with targeting micelles appeared to have higher signals than the non-targeting groups. Conclusion: LLY13 has promising in vitro and in vivo targeting activity against OSCC. In addition, LLY13 is also able to penetrate into cancer cells via endocytosis. Initial study indicated that alpha3 integrin might partially be the corresponding receptor involved

Endogenous Noxa Determines the Strong Proapoptotic Synergism of the BH3-Mimetic ABT-737 with Chemotherapeutic Agents in Human Melanoma Cells12

PubMed Central

Weber, Arnim; Kirejczyk, Zofia; Potthoff, Stephanie; Ploner, Christian; Häcker, Georg

2009-01-01

Human melanoma cells are very resistant to treatment with chemotherapeutic agents, and melanoma shows poor response to chemotherapeutic therapy. We describe a strong synergistic proapoptotic effect of the Bcl-2 family inhibitor ABT-737 and the standard antimelanoma drugs, namely, dacarbazine and fotemustine, and the experimental agent, imiquimod. Experiments with human melanoma cells, keratinocytes, and embryonic fibroblasts showed that all three agents activated the mitochondrial apoptosis pathway. ABT-737 on its own was ineffective in melanoma cells unless Mcl-1 was experimentally downregulated. However, ABT-737 strongly enhanced the proapoptotic activity of the chemotherapeutic drugs. Whereas cell death induction by all three agents involved the activity of both BH3-only proteins, Bim and Noxa, the combination with ABT-737 overcame the requirement for Bim. However, the synergism between ABT-737 and imiquimod or dacarbazine required endogenous Noxa, as demonstrated by experiments with Noxa-specific RNAi. Surprisingly, although Bim was activated, it was unable to replace Noxa. Studies of mitochondrial cytochrome c release using BH3 peptides confirmed that a main effect of dacarbazine, fotemustine, and imiquimod was to neutralize Mcl-1, thereby sensitizing mitochondria to the inhibition of other Bcl-2 family members through ABT-737. ABT-737 is thus a promising agent for combination therapy for human melanoma. Importantly, the efficacy of this therapy depends on endogenous Noxa, and the ability of chemotherapeutic drugs to activate Noxa may be a valuable predictor of their synergism with Bcl-2-targeting drugs. PMID:19412422

Inhibition of tomato (Solanum lycopersicum L.) root growth by cyanamide is not always accompanied with enhancement of ROS production.

PubMed

Soltys, Dorota; Gniazdowska, Agnieszka; Bogatek, Renata

2013-05-01

Mode of action of allelochemicals in target plants is currently widely studied. Cyanamide is one of the newly discovered allelochemical, biosynthesized in hairy vetch. Recently, it has been recognized that cyanamide is plant growth inhibitor, which affects mitosis in root tip cells and causes,e.g., disorder in phytohormonal balance. We also demonstrated that CA may act as oxidative stress agent but it strictly depends on plant species, exposure time and doses. Roots of tomato seedling treated with water solution of 1.2 mM cyanamide did not exhibit elevated reactive oxygen species concentration during the whole culture period.

Rapid screening of potential autophagic inductor agents using mammalian cell lines.

PubMed

Martins, Waleska K; Severino, Divinomar; Souza, Cleidiane; Stolf, Beatriz S; Baptista, Maurício S

2013-06-01

Recent progress in understanding the molecular basis of autophagy has demonstrated its importance in several areas of human health. Affordable screening techniques with higher sensitivity and specificity to identify autophagy are, however, needed to move the field forward. In fact, only laborious and/or expensive methodologies such as electron microscopy, dye-staining of autophagic vesicles, and LC3-II immunoblotting or immunoassaying are available for autophagy identification. Aiming to fulfill this technical gap, we describe here the association of three widely used assays to determine cell viability - Crystal Violet staining (CVS), 3-[4, 5-dimethylthiaolyl]-2, 5-diphenyl-tetrazolium bromide (MTT) reduction, and neutral red uptake (NRU) - to predict autophagic cell death in vitro. The conceptual framework of the method is the superior uptake of NR in cells engaging in autophagy. NRU was then weighted by the average of MTT reduction and CVS allowing the calculation of autophagic arbitrary units (AAU), a numeric variable that correlated specifically with the autophagic cell death. The proposed strategy is very useful for drug discovery, allowing the investigation of potential autophagic inductor agents through a rapid screening using mammalian cell lines B16-F10, HaCaT, HeLa, MES-SA, and MES-SA/Dx5 in a unique single microplate. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

1,2-Bis(methylsulfonyl)-1-(2-chloroethyl)-2-[[1-(4-nitrophenyl)ethoxy]carbonyl]hydrazine: An anticancer agent targeting hypoxic cells

PubMed Central

Seow, Helen A.; Penketh, Philip G.; Shyam, Krishnamurthy; Rockwell, Sara; Sartorelli, Alan C.

2005-01-01

To target malignant cells residing in hypoxic regions of solid tumors, we have designed and synthesized prodrugs generating the cytotoxic alkylating species 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine (90CE) after bioreductive activation. We postulate that one of these agents, 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[[1-(4-nitrophenyl)ethoxy]carbonyl]hydrazine (KS119), requires enzymatic nitro reduction to produce 90CE, whereas another agent, 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[(4-nitrobenzyloxy)carbonyl]hydrazine (PNBC), can also be activated by nucleophilic attack by thiols such as glutathione (GSH)/GST. We demonstrated that these agents selectively kill hypoxic EMT6 mouse mammary carcinoma and CHO cells. In hypoxia, 50 μM KS119 produced 5 logs of kill of EMT6 cells without discernable cytotoxicity in air; similar effects were observed with CHO cells. PNBC was less efficacious against hypoxic tumor cells and also had some toxicity to aerobic cells, presumably because of GST/thiol activation, making PNBC less interesting as a selective hypoxic-cell cytotoxin. BALB/c mice with established EMT6 solid tumors were used to demonstrate that KS119 could reach and kill hypoxic cells in solid tumors. To gain information on bioreductive enzymes involved in the activation of KS119, cytotoxicity was measured in CHO cell lines overexpressing NADH:cytochrome b5 reductase (NBR), NADPH:cytochrome P450 reductase (NPR), or NAD(P)H: quinone oxidoreductase 1 (NQO1). Increased cytotoxicity occurred in cells overexpressing NBR and NPR, whereas overexpressed NQO1 had no effect. These findings were supported by enzymatic studies using purified NPR and xanthine oxidase to activate KS119. KS119 has significant potential as a hypoxia-selective tumor-cell cytotoxin and is unlikely to cause major toxicity to well oxygenated normal tissues. PMID:15964988

The cell labeling efficacy, cytotoxicity and relaxivity of copper-activated MRI/PET imaging contrast agents.

PubMed

Patel, Daksha; Kell, Arnold; Simard, Benoit; Xiang, Bo; Lin, Hung Yu; Tian, Ganghong

2011-02-01

A new class of nanoparticle-based dual-modality positron emission tomography/magnetic resonance imaging (PET/MRI) contrast agents has been developed. The probe consists of a superparamagnetic iron oxide (SPIO) or manganese oxide core coated with 3,4-dihydroxy-D,L-phenylalanine (DL-DOPA). The chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) was conjugated to DOPA termini. The DOTA modified nanoparticles allow chelation of copper for PET imaging. These surface functionalized nanoparticle-based probes have been characterized by various analytical techniques. The cell-labeling efficacy, cytotoxicity and relaxivity of these nanoparticles have been evaluated and compared with the same properties of one of the most commonly utilized MRI contrast agents, Feridex(®). Evidently, this new nanoparticle has a great potential for use in cell tracking with MRI and PET in the absence of transfecting agent. It is noteworthy that there is a sharp increase in r(2) relaxivity of these nanoparticles on coordination with Cu(2+) ions. Thus these iron oxide nanoparticles can also be explored as the smart magnetic resonance (MR) sensor for the detection of micromolar changes in copper concentration for neurodegenerative diseases such as Alzheimer's disease, Menkes and Wilson's diseases, amyotrophic lateral sclerosis and prion diseases. Crown Copyright © 2010. Published by Elsevier Ltd. All rights reserved.

Loss of Nek11 Prevents G2/M Arrest and Promotes Cell Death in HCT116 Colorectal Cancer Cells Exposed to Therapeutic DNA Damaging Agents

PubMed Central

Sabir, Sarah R.; Sahota, Navdeep K.; Jones, George D. D.; Fry, Andrew M.

2015-01-01

The Nek11 kinase is a potential mediator of the DNA damage response whose expression is upregulated in early stage colorectal cancers (CRCs). Here, using RNAi-mediated depletion, we examined the role of Nek11 in HCT116 WT and p53-null CRC cells exposed to ionizing radiation (IR) or the chemotherapeutic drug, irinotecan. We demonstrate that depletion of Nek11 prevents the G2/M arrest induced by these genotoxic agents and promotes p53-dependent apoptosis both in the presence and absence of DNA damage. Interestingly, Nek11 depletion also led to long-term loss of cell viability that was independent of p53 and exacerbated following IR exposure. CRC cells express four splice variants of Nek11 (L/S/C/D). These are predominantly cytoplasmic, but undergo nucleocytoplasmic shuttling mediated through adjacent nuclear import and export signals in the C-terminal non-catalytic domain. In HCT116 cells, Nek11S in particular has an important role in the DNA damage response. These data provide strong evidence that Nek11 contributes to the response of CRC cells to genotoxic agents and is essential for survival either with or without exposure to DNA damage. PMID:26501353

Modulation of Regulatory T Cell Activity by TNF Receptor Type II-Targeting Pharmacological Agents

PubMed Central

Zou, Huimin; Li, Ruixin; Hu, Hao; Hu, Yuanjia; Chen, Xin

2018-01-01

There is now compelling evidence that tumor necrosis factor (TNF)–TNF receptor type II (TNFR2) interaction plays a decisive role in the activation, expansion, and phenotypical stability of suppressive CD4+Foxp3+ regulatory T cells (Tregs). In an effort to translate this basic research finding into a therapeutic benefit, a number of agonistic or antagonistic TNFR2-targeting biological agents with the capacity to activate or inhibit Treg activity have been developed and studied. Recent studies also show that thalidomide analogs, cyclophosphamide, and other small molecules are able to act on TNFR2, resulting in the elimination of TNFR2-expressing Tregs. In contrast, pharmacological agents, such as vitamin D3 and adalimumab, were reported to induce the expansion of Tregs by promoting the interaction of transmembrane TNF (tmTNF) with TNFR2. These studies clearly show that TNFR2-targeting pharmacological agents represent an effective approach to modulating the function of Tregs and thus may be useful in the treatment of major human diseases such as autoimmune disorders, graft-versus-host disease (GVHD), and cancer. In this review, we will summarize and discuss the latest progress in the study of TNFR2-targeting pharmacological agents and their therapeutic potential based on upregulation or downregulation of Treg activity. PMID:29632537

Rituximab in Treating Patients Undergoing Donor Peripheral Blood Stem Cell Transplant for Relapsed or Refractory B-cell Lymphoma

ClinicalTrials.gov

2017-12-05

B-cell Adult Acute Lymphoblastic Leukemia; B-cell Childhood Acute Lymphoblastic Leukemia; B-cell Chronic Lymphocytic Leukemia; Childhood Burkitt Lymphoma; Childhood Diffuse Large Cell Lymphoma; Childhood Immunoblastic Large Cell Lymphoma; Cutaneous B-cell Non-Hodgkin Lymphoma; Extranodal Marginal Zone B-cell Lymphoma of Mucosa-associated Lymphoid Tissue; Intraocular Lymphoma; Nodal Marginal Zone B-cell Lymphoma; Post-transplant Lymphoproliferative Disorder; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Adult Burkitt Lymphoma; Recurrent Adult Diffuse Large Cell Lymphoma; Recurrent Adult Diffuse Mixed Cell Lymphoma; Recurrent Adult Diffuse Small Cleaved Cell Lymphoma; Recurrent Adult Grade III Lymphomatoid Granulomatosis; Recurrent Adult Hodgkin Lymphoma; Recurrent Adult Immunoblastic Large Cell Lymphoma; Recurrent Adult Lymphoblastic Lymphoma; Recurrent Childhood Acute Lymphoblastic Leukemia; Recurrent Childhood Grade III Lymphomatoid Granulomatosis; Recurrent Childhood Large Cell Lymphoma; Recurrent Childhood Lymphoblastic Lymphoma; Recurrent Childhood Small Noncleaved Cell Lymphoma; Recurrent Grade 1 Follicular Lymphoma; Recurrent Grade 2 Follicular Lymphoma; Recurrent Grade 3 Follicular Lymphoma; Recurrent Mantle Cell Lymphoma; Recurrent Marginal Zone Lymphoma; Recurrent Small Lymphocytic Lymphoma; Recurrent/Refractory Childhood Hodgkin Lymphoma; Refractory Chronic Lymphocytic Leukemia; Refractory Hairy Cell Leukemia; Small Intestine Lymphoma; Splenic Marginal Zone Lymphoma; Testicular Lymphoma; Waldenström Macroglobulinemia

DOT1L inhibitor EPZ-5676 displays synergistic antiproliferative activity in combination with standard of care drugs and hypomethylating agents in MLL-rearranged leukemia cells.

PubMed

Klaus, Christine R; Iwanowicz, Dorothy; Johnston, Danielle; Campbell, Carly A; Smith, Jesse J; Moyer, Mikel P; Copeland, Robert A; Olhava, Edward J; Scott, Margaret Porter; Pollock, Roy M; Daigle, Scott R; Raimondi, Alejandra

2014-09-01

EPZ-5676 [(2R,3R,4S,5R)-2-(6-amino-9H-purin-9-yl)-5-((((1r,3S)-3-(2-(5-(tert-butyl)-1H-benzo[d]imidazol-2-yl)ethyl)cyclobutyl)(isopropyl)amino)methyl)tetrahydrofuran-3,4-diol], a small-molecule inhibitor of the protein methyltransferase DOT1L, is currently under clinical investigation for acute leukemias bearing MLL-rearrangements (MLL-r). In this study, we evaluated EPZ-5676 in combination with standard of care (SOC) agents for acute leukemias as well as other chromatin-modifying drugs in cellular assays with three human acute leukemia cell lines: MOLM-13 (MLL-AF9), MV4-11 (MLL-AF4), and SKM-1 (non-MLL-r). Studies were performed to evaluate the antiproliferative effects of EPZ-5676 combinations in a cotreatment model in which the second agent was added simultaneously with EPZ-5676 at the beginning of the assay, or in a pretreatment model in which cells were incubated for several days in the presence of EPZ-5676 prior to the addition of the second agent. EPZ-5676 was found to act synergistically with the acute myeloid leukemia (AML) SOC agents cytarabine or daunorubicin in MOLM-13 and MV4-11 MLL-r cell lines. EPZ-5676 is selective for MLL-r cell lines as demonstrated by its lack of effect either alone or in combination in the nonrearranged SKM-1 cell line. In MLL-r cells, the combination benefit was observed even when EPZ-5676 was washed out prior to the addition of the chemotherapeutic agents, suggesting that EPZ-5676 sets up a durable, altered chromatin state that enhances the chemotherapeutic effects. Our evaluation of EPZ-5676 in conjunction with other chromatin-modifying drugs also revealed a consistent combination benefit, including synergy with DNA hypomethylating agents. These results indicate that EPZ-5676 is highly efficacious as a single agent and synergistically acts with other chemotherapeutics, including AML SOC drugs and DNA hypomethylating agents in MLL-r cells. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

Monitoring and Management of Toxicities of Novel B Cell Signaling Agents.

PubMed

Rhodes, Joanna; Mato, Anthony; Sharman, Jeff P

2018-04-11

B cell signaling agents, including ibrutinib, idelalisib, and the BCL-2 inhibitor venetoclax have become an integral part of therapy for patients with non-Hodgkin's lymphomas. The toxicity profiles of these medications is distinct from chemoimmunotherapy. Here, we will review the mechanism of action of these drugs, their efficacy, and toxicity management. Ibrutinib use is associated with increased risk of atrial fibrillation and bleeding which can be managed using dose interruptions and modifications. Patients on idelalisib require close clinical and frequent laboratory monitoring, particularly of liver function tests to ensure there are no serious adverse events. Monitoring for infections is important in patients on both idelalisib and ibrutinib. Venetoclax requires close clinical and laboratory monitoring to prevent significant tumor lysis. Targeted B cell receptor therapies each have unique side effect profiles which require careful clinical monitoring. As we continue to use these therapies, optimal management strategies will continue to be elucidated.

Adsorption of hairy particles with mobile ligands: Molecular dynamics and density functional study

NASA Astrophysics Data System (ADS)

Borówko, M.; Sokołowski, S.; Staszewski, T.; Pizio, O.

2018-01-01

We study models of hairy nanoparticles in contact with a hard wall. Each particle is built of a spherical core with a number of ligands attached to it and each ligand is composed of several spherical, tangentially jointed segments. The number of segments is the same for all ligands. Particular models differ by the numbers of ligands and of segments per ligand, but the total number of segments is constant. Moreover, our model assumes that the ligands are tethered to the core in such a manner that they can "slide" over the core surface. Using molecular dynamics simulations we investigate the differences in the structure of a system close to the wall. In order to characterize the distribution of the ligands around the core, we have calculated the end-to-end distances of the ligands and the lengths and orientation of the mass dipoles. Additionally, we also employed a density functional approach to obtain the density profiles. We have found that if the number of ligands is not too high, the proposed version of the theory is capable to predict the structure of the system with a reasonable accuracy.

Agent-Based Computational Modeling of Cell Culture: Understanding Dosimetry In Vitro as Part of In Vitro to In Vivo Extrapolation

EPA Science Inventory

Quantitative characterization of cellular dose in vitro is needed for alignment of doses in vitro and in vivo. We used the agent-based software, CompuCell3D (CC3D), to provide a stochastic description of cell growth in culture. The model was configured so that isolated cells assu...

Correction of the Abnormal Trafficking of Primary Myelofibrosis CD34+ Cells by Treatment with Chromatin Modifying Agents

PubMed Central

Wang, Xiaoli; Zhang, Wei; Ishii, Takefumi; Sozer, Selcuk; Wang, Jiapeng; Xu, Mingjiang; Hoffman, Ronald

2011-01-01

The abnormal trafficking of CD34+ cells is a unique characteristic of primary myelofibrosis (PMF). We have further studied the behavior of PMF CD34+ cells by examining their homing to the marrow and the spleens of NOD/SCID mice. Following the infusion of PMF and normal G-CSF mobilized peripheral blood (mPB) CD34+ cells into NOD/SCID mice, reduced numbers of PMF CD34+ cells and CFU-GM as compared to mPB were detected in the marrow of these mice while similar numbers of PMF and mPB CD34+ cells and CFU-GM homed to their spleens. The abnormal homing of PMF CD34+ cells was associated with reduced expression of CXCR4, but was not related to the presence of JAK2V617F. The sequential treatment of PMF CD34+ cell with the chromatin modifying agents, 5-aza-2'-deoxycytidine (5azaD) and trichostatin A (TSA) but not treatment with small molecule inhibitors of JAK2 resulted in the generation of increased numbers of CD34+CXCR4+ cells which was accompanied by enhanced homing of PMF CD34+ cells to the marrow but not the spleens of NOD/SCID mice. Following 5azaD/TSA treatment JAK2V617F negative PMF hematopoietic progenitor cells preferentially homed to the marrow but not the spleens of recipient mice. Our data suggest that PMF CD34+ cells are characterized by a reduced ability to home to the marrow but not the spleens of NOD/SCID mice and that this homing defect can be corrected by sequential treatment with chromatin modifying agents. PMID:19752087

Functionalization of calcium carbonate microparticles as a combined sensor and transport system for active agents in cells.

PubMed

Reibetanz, Uta; Chen, Min Hui Averil; Mutukumaraswamy, Shaillender; Liaw, Zi Yen; Oh, Bernice Hui Lin; Donath, Edwin; Neu, Björn

2011-01-01

In recent years colloidal particles and capsules, layer-by-layer (LbL) coated with biocompatible polyelectrolytes, have received much attention as drug-delivery systems. In this study an LbL-assembled, biopolymer-based multilayer system was established as a combined transporter and sensor for monitoring intracellular degradation and processing. CaCO(3) cores were functionalized with fluorescein isothiocyanatelabelled poly(allylamine hydrochloride) (FITC-PAH). This pH-sensitive fluorescent dye allows identifying the location of these LbL-coated particles in cell compartments of different pH, like the endo-lysosome and cytoplasm. The labelled core was then coated with consecutive layers of protamine (PRM) and dextran sulfate (DXS). Finally, plasmid DNA (pEGFP-C1) as a reporter agent for drug release in the cytoplasm was integrated into the biocompatible and degradable PRM/DXS multilayer. The system was tested regarding its long-term stability and interaction with HEK 293T/17 cells. These multifunctional microparticles allow the simultaneous investigation of particle localization and processing within cells, and should thus provide a valuable tool for studying and improving the controlled LbL-based release of active agents into cells. © Koninklijke Brill NV, Leiden, 2011

Contrast agent-free sonoporation: The use of an ultrasonic standing wave microfluidic system for the delivery of pharmaceutical agents

PubMed Central

Carugo, Dario; Ankrett, Dyan N.; Glynne-Jones, Peter; Capretto, Lorenzo; Boltryk, Rosemary J.; Zhang, Xunli; Townsend, Paul A.; Hill, Martyn

2011-01-01

Sonoporation is a useful biophysical mechanism for facilitating the transmembrane delivery of therapeutic agents from the extracellular to the intracellular milieu. Conventionally, sonoporation is carried out in the presence of ultrasound contrast agents, which are known to greatly enhance transient poration of biological cell membranes. However, in vivo contrast agents have been observed to induce capillary rupture and haemorrhage due to endothelial cell damage and to greatly increase the potential for cell lysis in vitro. Here, we demonstrate sonoporation of cardiac myoblasts in the absence of contrast agent (CA-free sonoporation) using a low-cost ultrasound-microfluidic device. Within this device an ultrasonic standing wave was generated, allowing control over the position of the cells and the strength of the acoustic radiation forces. Real-time single-cell analysis and retrospective post-sonication analysis of insonated cardiac myoblasts showed that CA-free sonoporation induced transmembrane transfer of fluorescent probes (CMFDA and FITC-dextran) and that different mechanisms potentially contribute to membrane poration in the presence of an ultrasonic wave. Additionally, to the best of our knowledge, we have shown for the first time that sonoporation induces increased cell cytotoxicity as a consequence of CA-free ultrasound-facilitated uptake of pharmaceutical agents (doxorubicin, luteolin, and apigenin). The US-microfluidic device designed here provides an in vitro alternative to expensive and controversial in vivo models used for early stage drug discovery, and drug delivery programs and toxicity measurements. PMID:22662060