scholarly journals Producción y evaluación inmunoquímica de extractos alergénicos del ácaro del polvo doméstico Blomia tropicalis (Acari: Echymyopodidae)

2018 ◽  
Vol 21 ◽  
Author(s):  
Luis Acuña-Cantillo ◽  
Dary Luz Mendoza-Meza ◽  
Gloria Garavito ◽  
Ana Sofía Moreno-Woo ◽  
Eduardo Egea-Bermejo
Keyword(s):  
Western Blot ◽  
Blomia Tropicalis ◽  
Sds Page

Los alérgenos de Blomia tropicalis son un factor de riesgo para el desarrollo de alergias en países tropicales. Los extractos alergénicos son reactantes valiosos para el diagnóstico de la hipersensibilidad alérgica a los ácaros y su control biológico. El objetivo fue evaluar las propiedades inmunoquímicas de extractos de B. tropicalis producidos desde cultivos in vitro. Este trabajo no tuvo como propósito producir extractos de aplicación clínica. Los ejemplares de B. tropicalis se establecieron a partir de ácaros aislados del polvo doméstico en la ciudad de Barranquilla, Colombia. Los ácaros libres de medio de cultivo se usaron para la obtención de los extractos. El rendimiento de la extracción se determinó mediante ensayo Bradford y el perfil electroforético por SDS-PAGE. La presencia de enzimas hidrolasas se determinó por Api® Zym y la inmunogenicidad se evaluó mediante Western blot donde se identificaron proteínas de unión a IgE sérica. La producción máxima de ácaros en los cultivos fue de 0,9 g/300 g medio. El rendimiento de extracción de proteínas fue de 3,61 % (± 0,42). Todos los extractos presentaron actividad enzimática tripsina, alfa-amilasa y quimotripsina. Western blot confirmó la presencia de proteínas de unión a IgE sérica. Presentamos un método sencillo para obtener extractos del ácaro B. tropicalis con actividad enzimática y alergénica.

scholarly journals Σύνθεση και μελέτη της δομής και της βιοδραστικότητας οργανοκασσιτερικών ενώσεων με αρυλιμινο υποκατεστημένα βενζοϊκά οξέα

2015 ◽  
Author(s):  
Muvudji Cephas Mwanasaka
Keyword(s):  
Western Blot ◽  
1H Nmr ◽  
13C Nmr ◽  
Sds Page ◽  
Ft Ir

Στα πλαίσια της παρούσας διδακτορικής διατριβής πραγματοποιήθηκε μια διεπιστημονική ερευνητική εργασία, η οποία είχε ως κεντρική ιδέα τη χημεία και τη βιοδραστικότητα των οργανοκασσιτερικών ενώσεων. Για την επίτευξη της μελέτης αυτής, η ερευνητική εργασία κινήθηκε σε τρεις κεντρικούς άξονες: σχεδιασμός και σύνθεση, ανάλυση και ταυτοποίηση και, τέλος, in vitro μελέτη βιολογικών δράσεων. Αρχικά σχεδιάστηκαν και συντέθηκαν ιμίνες (βάσεις Schiff) παράγωγα του βενζοϊκού οξέος, από τη συμπύκνωση του 3- και του 4-αμινοβενζοϊκου οξέος με π-υποκατεστημένες (Η, ΝΟ2, Cl, MeO, Me2N) βενζαλδεΰδες, καθώς και από τη συμπύκνωση των π-υποκατεστημένων (Η, ΝΟ2, Cl, MeO, Me2N) ανιλίνων με π-φορμυλοβενζοϊκό οξύ. Από τις αντιδράσεις αυτές συντέθηκαν βιολογικά ενεργά αρυλιμηνο υποκατεστημένα βενζοϊκά οξέα, τα οποία χρησιμοποιήθηκαν στη συνέχεια της συνθετικής διαδικασίας ως ligands (L) για τη σύνθεση νέων τριοργανοκασσιτερικών εστέρων γενικού τύπου R3SnL με R= Μe και Ph. Η ταυτοποίηση και ο χαρακτηρισμός των νέων αυτών ligands και των συμπλόκων που συντέθηκαν πραγματοποιήθηκε με συνδυασμό αναλυτικών τεχνικών: φασματοσκοπικές τεχνικές (FT-IR, 1H-NMR, 13C-NMR, UV-vis), στοιχειακή ανάλυση και την κρυσταλλογραφία ακτίνων Χ. Επόμενο στάδιο ήταν η in vitro μελέτη βιοδραστικότητας των ενώσεων αυτών η οποία, χωρίστηκε σε δύο σκέλη. Το ένα βιολογικό σκέλος κινήθηκε σε κυτταρικό επίπεδο, όπου αξιολογήθηκε πρώτα η in vitro αποπτωτική δράση σε τέσσερεις καρκινικές κυτταρικές σειρές, ΗepG2, HeLa, K562 και N2a. Για το σκοπό αυτό χρησιμοποιήθηκε η δοκιμή ΜΤΤ (ΜΤΤ assay), η οποία εκτιμά την επίδραση στη βιωσιμότητα των κυττάρων μετά από έκθεσή τους σε διάφορες συγκεντρώσεις των ουσιών. Στη συνέχεια έγιναν νευροκυτταροτοξικές μελέτες, εκτιμώντας την ενδεχόμενη in vitro αναπτυξιακή νευροτοξικότητα. Για το σκοπό αυτό αξιολογήθηκε η επίδραση των υπό μελέτη ουσιών στη διαδικασία ανάπτυξης αξονικών προεκβολών σε υπό διαφοροποίηση Ν2a κύτταρα, μετά από έκθεση σε υποκυτταροτοξικές συγκεντρώσεις των υπό μελέτη ουσιών. Το άλλο βιολογικό σκέλος κινήθηκε σε μοριακό επίπεδο, στοχεύοντας στην εκτίμηση της επίδρασης των υπό μελέτη ουσιών σε συγκεκριμένους βιοχημικούς παράγοντες με σκοπό τη διαλεύκανση των βιοχημικών μηχανισμών δράσεών τους. Εκτιμήθηκε η δράση τους σε συγκεκριμένες πρωτεΐνες του νευροκυτταρικού σκελετού, χρησιμοποιώντας SDS-PAGE και ανοσοχημική τεχνική ανάλυσης Western blot. Επιπλέον, έγινε εκτίμηση της in vitro ανασταλτικής δράσης τους στη λιποξυγονάση, ένζυμο που είναι γνωστό για το ρόλο του στην παραγωγή ελευθέρων ριζών στη φλεγμονή. Τέλος, εκτιμήθηκε η ενδεχόμενη in vitro αντιοξειδωτική δράση τους, αξιολογώντας την ικανότητά τους στην αναστολή της επαγόμενης από το ΑΑΡΗ λιπιδικής υπεροξείδωσης, καθώς και την ικανότητα αλληλεπίδρασής τους με τη σταθερή ρίζα DPPH. Από τα αποτελέσματα που προέκυψαν προσπαθήσαμε να βγάλουμε συμπεράσματα για το κατά πόσο η δομή των καρβοξυλικών οξέων που χρησιμοποιήθηκαν ως Ligands επηρεάζει τόσο τον τρόπο συναρμογής της καρβοξυλικής ομάδας με το τριοργανοκασσιτερικό κατιόν, όσο και τη βιολογική δράση των υπό μελέτη τριοργανοκασσιτερικών εστέρων στις προαναφερόμενες μελέτες.

scholarly journals Expression of Grapevine leafroll-associated virus 3 coat protein gene in Escherichia coli and production of polyclonal antibodies

2007 ◽  
Vol 32 (6) ◽  
pp. 496-500 ◽  
Author(s):  
Thor V.M. Fajardo ◽  
Danielle R. Barros ◽  
Osmar Nickel ◽  
Gilmar B. Kuhn ◽  
F. Murilo Zerbini
Keyword(s):  
Escherichia Coli ◽  
Coat Protein ◽  
Western Blot ◽  
Coat Protein Gene ◽  
Protein Gene ◽  
Polyclonal Antibodies ◽  
Yield And Quality ◽  
Sds Page ◽  
Grapevine Leaves

Grapevine leafroll-associated virus 3 (GLRaV-3), the main viral species of the grapevine leafroll complex, causes yield and quality reduction in grapes (Vitis spp.). The coat protein gene was RT-PCR-amplified from total RNA extracted from infected grapevine leaves and the amplified fragment was cloned and completely sequenced. The fragment was subsequently subcloned into the pRSET-C expression vector. The recombinant plasmid was used to transform Escherichia coli BL21:DE3 and express the capsid protein. The coat protein, fused to a 6 His-tag, was purified by affinity chromatography using an Ni-NTA resin. The identity of the purified protein was confirmed by SDS-PAGE and Western blot. The in vitro-expressed protein was quantified and used for rabbit immunizations. The antiserum was shown to be sensitive and specific for the detection of GLRaV-3 in grapevine extracts in Western blot and DAS-ELISA assays, with no unspecific or heterologous reactions against other non-serologically related viruses being observed.

In vitro Aggregation Ability of Five Commercially Available Aβ42 Peptides

2021 ◽  
Vol 18 ◽  
Author(s):  
Zhaoji Lv ◽  
Xi Du ◽  
Zhongsheng Chen ◽  
Fengjuan Liu ◽  
Rong Zhang ◽  
...  
Keyword(s):  
Western Blot ◽  
Amyloid Β ◽  
Cell Counting ◽  
Basic Material ◽  
Aggregation Kinetics ◽  
Basic Study ◽  
Sds Page ◽  
Sigmoidal Kinetics ◽  
High Cytotoxicity

Background: As the most basic material, synthetic human Amyloid-β (1-42) (Aβ42) pep- tides from different manufacturers have been widely used. Their aggregation ability is vital to the reliability, repeatability and comparability of studies on Aβ42 physiology and pathology. However, it has not been evaluated and compared. Objectives: To analyze the consistency of the aggregation ability of 5 commercially available Aβ42 peptides. Methods: 5 Aβ42 peptides represented as A, B, C, D and E were pretreated by HFIP. The pretreated Aβ42 peptides were dissolved in Thioflavin T (ThT) solution, and their aggregation kinetics was monitored for 30 h with the aggregation kinetics test. Meanwhile, the pretreated peptides were ag- gregated in phosphate buffered saline. After aggregated for 12 h, they were detected by methods of ThT fluorescence, far-UV circular dichroism (CD), SDS-PAGE, western blot, and transmission electron microscopy (TEM), respectively. After aggregation for 8 h and 12 h, their cytotoxicity to SH-SY5Y cells was further evaluated using Cell Counting Kit-8. Results: For aggregation kinetics, peptides A, C and E remained low level curves, while peptides B and D presented typical sigmoidal kinetics curves. In CD measurement, the aggregates of pep- tides B and D showed relatively high negative CD peaks with the height of -8.09 mdeg and -14.37 mdeg, while the height of peptides A, C and E was -1.04, -3.55, and -3.88. In ThT assay, relative fluorescence intensity of the aggregates of peptides B and D were 7.79 and 8.82, higher than 1.19, 1.71, and 2.70 of peptide A, C and E, respectively. In SDS-PAGE, all aggregates contained monomers and eleven polymers. Moreover, peptide B-E presented a trapezoidal distribution from dimers to trimers, and peptide A aggregated to dimers. By western blot, the bands of monomers re- mained in all aggregates. Furthermore, peptides B and D aggregated to dimers and trimers, pep- tides A and C only aggregated to dimers, and peptide E showed a strong band of trimers. By TEM, protofibrils were observed only in peptide B, while substantial spherical aggregates were formed in other peptides. Additionally, peptides B, D and E exhibited higher cytotoxicity after being aggregat- ed for 8 h, whereas peptides A, B and D presented relatively high cytotoxicity after 12-hour aggre- gation. Conclusion: Commercially available Aβ42 peptides showed obvious differences in aggregation abil- ity, which should arouse enough attention in the field of basic study related to Aβ42. The aggrega- tion ability evaluation with the various assay methods has some discrepancies, and it is highly ur- gent to establish a reasonable and uniform measurement strategy.

Comparative evaluation of PAR1, GPIb-IX-V, and integrin αIIbβ3 levels in cord and adult platelets

2010 ◽  
Vol 30 (S 01) ◽  
pp. S164-S167 ◽  
Author(s):  
A. Schlagenhauf ◽  
S. Schweintzger ◽  
R. Birner-Grünberger ◽  
B. Leschnik ◽  
W. Muntean
Keyword(s):  
Statistical Analysis ◽  
Western Blot ◽  
Cord Blood ◽  
Comparative Evaluation ◽  
Clot Formation ◽  
Thrombin Receptor ◽  
Lower Content ◽  
Integrin Αiibβ3 ◽  
Sds Page

Summary Background: Newborn platelets show hyposensitivity in vitro to many important agonists. However, sensitivity of platelets to these agonists is crucial for a functional clot formation. Nevertheless newborns have an excellent hemostasis. Hence, we examined levels of PAR1 thrombin receptor, GPIb-IX-V (CD42b), and Integrin αIIbβ3 in newborn and adult platelets using Western blot analysis.Materials, methods: Platelets of adult and cord blood were isolated, washed, and lysed. Protein samples were separated by SDS-PAGE and blotted on nitrocellulose membranes. Receptors were visualized using immunodetection and evaluated densitometrically. Statistical analysis was performed using SPSS 16.0. Results: We found significantly lower levels of PAR1-receptors and higher levels of CD42b in newborn platelets as compared to adult platelets. Levels of Integrin αIIbβ3 in newborn platelets were comparable to adult platelets. Conclusion: A lower content of PAR1-receptors explains very well the hyposensitivity of cord platelets to thrombin. Higher levels of CD42b may additionally support the effect of larger more adhesive multimeric vWF in newborn plasma.

scholarly journals Produção de antissoro policlonal utilizando a proteína capsidial recombinante do Rupestris stem pitting-associated virus

2010 ◽  
Vol 40 (11) ◽  
pp. 2385-2388 ◽  
Author(s):  
Marcos Fernando Basso ◽  
Thor Vinícius Martins Fajardo ◽  
Marcelo Eiras ◽  
Ricardo Antônio Ayub ◽  
Osmar Nickel
Keyword(s):  
Escherichia Coli ◽  
Western Blot ◽  
E Coli ◽  
Sds Page ◽  
Rupestris Stem Pitting ◽  
Stem Pitting

O Rupestris stem pitting-associated virus (RSPaV) é o agente causal das caneluras do lenho da videira. Este trabalho teve como objetivo produzir antissoro policlonal a partir da proteína capsidial (CP) recombinante do RSPaV e avaliar a sua especificidade e sensibilidade. O gene da CP do RSPaV, com 780pb, foi previamente caracterizado. Esse gene foi subclonado no sítio de restrição EcoRI, no vetor de expressão pRSET-B e o plasmídeo recombinante foi utilizado para induzir a expressão da CP em Escherichia coli. A CP, ligada a uma cauda de seis histidinas, foi purificada por meio de cromatografia de afinidade em coluna de Ni-NTA a partir do extrato de proteínas totais extraídas de E. coli. A identidade da proteína purificada foi confirmada em SDS-PAGE e Western blot, utilizando-se anticorpos comerciais contra a cauda de seis histidinas. A CP recombinante expressada in vitro apresentou massa molecular de cerca de 31kDa. A proteína purificada foi quantificada e 2,55mg foram utilizados para a imunização de um coelho. O antissoro policlonal obtido reagiu com diferentes isolados deste vírus, extraídos de videiras em ELISA indireto.

The Anticoagulant Properties of a Modified Form of Protein S

1988 ◽  
Vol 60 (02) ◽  
pp. 298-304 ◽  
Author(s):  
C A Mitchell ◽  
S M Kelemen ◽  
H H Salem
Keyword(s):  
Monoclonal Antibody ◽  
Anticoagulant Activity ◽  
Activated Protein C ◽  
Factor Xa ◽  
Protein S ◽  
Human Neutrophil Elastase ◽  
Factor X ◽  
Sds Page

SummaryProtein S (PS) is a vitamin K-dependent anticoagulant that acts as a cofactor to activated protein C (APC). To date PS has not been shown to possess anticoagulant activity in the absence of APC.In this study, we have developed monoclonal antibody to protein S and used to purify the protein to homogeneity from plasma. Affinity purified protein S (PSM), although identical to the conventionally purified protein as judged by SDS-PAGE, had significant anticoagulant activity in the absence of APC when measured in a factor Xa recalcification time. Using SDS-PAGE we have demonstrated that prothrombin cleavage by factor X awas inhibited in the presence of PSM. Kinetic analysis of the reaction revealed that PSM competitively inhibited factor X amediated cleavage of prothrombin. PS preincubated with the monoclonal antibody, acquired similar anticoagulant properties. These results suggest that the interaction of the monoclonal antibody with PS results in an alteration in the protein exposing sites that mediate the observed anticoagulant effect. Support that the protein was altered was derived from the observation that PSM was eight fold more sensitive to cleavage by thrombin and human neutrophil elastase than conventionally purified protein S.These observations suggest that PS can be modified in vitro to a protein with APC-independent anticoagulant activity and raise the possibility that a similar alteration could occur in vivo through the binding protein S to a cellular or plasma protein.

Isolation and Structural Charaderization of a Potent Inhibitor of Coagulation Factor Xa from the Leech Haementeria ghilianii

1989 ◽  
Vol 61 (03) ◽  
pp. 437-441 ◽  
Author(s):  
Cindra Condra ◽  
Elka Nutt ◽  
Christopher J Petroski ◽  
Ellen Simpson ◽  
P A Friedman ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Salivary Gland ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Western Blot ◽  
Potent Inhibitor ◽  
Factor Xa ◽  
Coagulation Factor ◽  
Sds Page ◽  
Bovine Factor

SummaryThe present work reports the discovery and charactenzation of an anticoagulant protein in the salivary gland of the giant bloodsucking leech, H. ghilianii, which is a specific and potent inhibitor of coagulation factor Xa. The inhibitor, purified to homogeneity, displayed subnanomolar inhibition of bovine factor Xa and had a molecular weight of approximately 15,000 as deduced by denaturing SDS-PAGE. The amino acid sequence of the first 43 residues of the H. ghilianii derived inhibitor displayed a striking homology to antistasin, the recently described subnanomolar inhibitor of factor Xa isolated from the Mexican leech, H. officinalis. Antisera prepared to antistasin cross-reacted with the H. ghilianii protein in Western Blot analysis. These data indicate that the giant Amazonian leech, H. ghilianii, and the smaller Mexican leech, H. officinalrs, have similar proteins which disrupt the normal hemostatic clotting mechanisms in their mammalian host’s blood.

In Vitro Stability of a Tissue-Type Plasminogen Activator Mutant, BM 06.022, in Human Plasma

1994 ◽  
Vol 72 (06) ◽  
pp. 906-911 ◽  
Author(s):  
D C Rijken ◽  
E Groeneveld ◽  
M M Barrett-Bergshoeff
Keyword(s):  
Plasminogen Activator ◽  
Half Life ◽  
Polyacrylamide Gel Electrophoresis ◽  
Tissue Type ◽  
Single Chain ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Sds Page ◽  
Chain Form

SummaryBM 06.022 is a non-glycosylated mutant of human tissue-type plasminogen activator (t-PA) comprising only the kringle-2 and proteinase domains. The in vivo half-life of BM 06.022 antigen is 4- to 5-fold longer than that of t-PA antigen. The in vitro half-life of the activity of BM 06.022 at therapeutic concentrations in plasma is shorter than that of t-PA. In this study the inactivation of BM 06.022 in plasma was further investigated.Varying concentrations of BM 06.022 were incubated in plasma for 0-150 min. Activity assays on serial samples showed a dose-dependent decline of BM 06.022 activity with a half-life from 72 min at 0.3 μg/ml to 38 min at 10 μg/ml. SDS-polyacrylamide gel electrophoresis (SDS-PAGE) followed by fibrin autography showed the generation of several BM 06.022-complexes. These complexes could be completely precipitated with antibodies against Cl-inactivator, α2-antiplasmin and α1-antitrypsin.During the incubation of BM 06.022 in plasma, plasmin was generated dose-dependently as revealed by varying degrees of a2-anti-plasmin consumption and fibrinogen degradation. SDS-PAGE and immunoblotting showed that single-chain BM 06.022 was rapidly (i. e. within 45 min) converted into its two-chain form at concentrations of 5 μg/ml BM 06.022 and higher.In conclusion, BM 06.022 at therapeutic concentrations in plasma was inactivated by Cl-inactivator, a2-antiplasmin and a j-antitrypsin. The half-life of the activity decreased at increasing BM 06.022 concentrations, probably as a result of the generation of two-chain BM 06.022 which may be inactivated faster than the single-chain form.

Western Blot Analyses of Prekallikrein and its Activation Products in Human Plasma

1991 ◽  
Vol 65 (04) ◽  
pp. 382-388 ◽  
Author(s):  
Dulce Veloso ◽  
Robert W Colman
Keyword(s):  
Complex Formation ◽  
Western Blot ◽  
Human Plasma ◽  
Contact System ◽  
Plasma Activation ◽  
Normal Plasma ◽  
Sds Page ◽  
Activation Products ◽  
Major Antigen ◽  
Formation Of Complexes

SummaryPrekallikrein (PK), a zymogen of the contact system, and its activation products, kallikrein (KAL), KAl-inhibitor complexes and fragments containing KAL epitope(s) have been detected in human plasma by immunoblotting with a monoclonal anti-human plasma PK antibody, MAb 13G1L. Detection of antigen-MAb 13G11 complexes with peroxidase-conjugated anti-IgG showed that the two variants of PK (85- and 88-kDa) are the only major antigen species in normal, non-activated plasma. Upon plasma activation with kaolin, the intensity of the PK bands decreased with formation of complexes of KAL with CL inhibitor (C1 INH) and α2-rrtzcroglobulin (α2M) identical to those formed by the purified proteins. Immunoblots of normal plasma showed good correlation between the PK detected and the amount of plasma assayed. Increasing amounts of KAL incubated with a constant volume of PK-deficient plasma showed increasing amounts of KAL and of KAL-C1 INH and KAL-α2M complexes. Complexes of KALantithrombin III (ATIII) and the ratio of KALα2M/ KAL-CL INH were higher in activated CL INH-deficient plasmas than in activated normal plasmas. Protein resolution by 3-12% gradient SDS-PAGE and epitope detection with [125I]MAb 13G11 showed four KALα2M species and a 45-kDa fragment(s) in both surface-activated normal plasma and complexes formed by purified KAL and α2M. Immunoblots of activated plasma also showed bands at the position of KALCL INH and KALATIII complexes. When α1-antitrypsin Pittsburgh (cα1-AT, Pitts) was added to plasma before activation, KAL-α1-ALPitts was the main complex. The non-activated normal plasma revealed only an overloaded PK band. This is the first report of an antibody that recognizes KAL epitope(s) in KAL-α2M, KALATIII and KALa1-α1Pitts complexes and in the 45-kDa fragment(s). Therefore, MAb 13G11 should be useful for studying the structure of these complexes as well as the mechanism of complex formation. In addition, immunoblotting with MAb 13G11 would allow detection of KAl-inhibitor complexes in patient plasmas as indicators of activation of the contact system.

Radiosensitivity of glioma to Gamma Knife treatment enhanced in vitro and in vivo by RNA interfering Ku70 that is mediated by a recombinant adenovirus

2010 ◽  
Vol 113 (Special_Supplement) ◽  
pp. 228-235 ◽  
Author(s):  
Qiang Jia ◽  
Yanhe Li ◽  
Desheng Xu ◽  
Zhenjiang Li ◽  
Zhiyuan Zhang ◽  
...  
Keyword(s):  
Western Blot ◽  
Gamma Knife ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Glioma Cells ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
C6 Cells

Object The authors sought to evaluate modification of the radiation response of C6 glioma cells in vitro and in vivo by inhibiting the expression of Ku70. To do so they investigated the effect of gene transfer involving a recombinant replication-defective adenovirus containing Ku70 short hairpin RNA (Ad-Ku70shRNA) combined with Gamma Knife treatment (GKT). Methods First, Ad-Ku70shRNA was transfected into C6 glioma cells and the expression of Ku70 was measured using Western blot analysis. In vitro, phenotypical changes in C6 cells, including proliferation, cell cycle modification, invasion ability, and apoptosis were evaluated using the MTT (3′(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide) assay, Western blot analysis, and cell flow cytometry. In vivo, parental C6 cells transfected with Ad-Ku70shRNA were implanted stereotactically into the right caudate nucleus in Sprague-Dawley rats. After GKS, apoptosis was analyzed using the TUNEL (terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick-end labeling) method. The inhibitory effects on growth and invasion that were induced by expression of proliferating cell nuclear antigen and matrix metalloproteinase–9 were determined using immunohistochemical analyses. Results The expression of Ku70 was clearly inhibited in C6 cells after transfection with Ad-Ku70shRNA. In vitro following transfection, the C6 cells showed improved responses to GKT, including suppression of proliferation and invasion as well as an increased apoptosis index. In vivo following transfection of Ad-Ku70shRNA, the therapeutic efficacy of GKT in rats with C6 gliomas was greatly enhanced and survival times in these animals were prolonged. Conclusions Our data support the potential for downregulation of Ku70 expression in enhancing the radiosensitivity of gliomas. The findings of our study indicate that targeted gene therapy–mediated inactivation of Ku70 may represent a promising strategy in improving the radioresponsiveness of gliomas to GKT.

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