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Rescue of cardiac failing and remodelling by inhibition of protein phosphatase 1γ is associated with suppression of the alternative splicing factor-mediated splicing of Ca2+/calmodulin-dependent protein kinase δ.

Our previous studies showed that protein phosphatase 1γ (PP1γ) exacerbates cardiomyocyte apoptosis through promotion of Ca(2+)/calmodulin-dependent protein kinase δ (CaMKIIδ) splicing. Here we determine the role of PP1γ in abdominal aorta constriction-induced hypertrophy and remodelling in rat hearts. Systolic blood pressure and echocardiographic measurements were used to evaluate the model of cardiac hypertrophy. Sirius red staining and invasive haemodynamic/cardiac index measurements were used to evaluate the effects of PP1γ or inhibitor 1 of PP1 transfection. Western blot, reverse transcrip…

Japanese encephalitis virus induces matrix metalloproteinase-9 in rat brain astrocytes via NF-κB signalling dependent on MAPKs and reactive oxygen species.

BACKGROUND AND PURPOSE: Japanese encephalitis virus (JEV) is a member of the family Flaviviridae and JEV infection is a major cause of acute … factor NF-κB in these responses was investigated by using selective pharmacological inhibitors and transfection with appropriate siRNAs. KEY RESULTS: JEV induced the expression of the pro-form of MMP-9 in rat brain astrocytes (RBA-1 cells). In RBA-1 cells, JEV induced MMP-9 expression and promoter activity, which was inhibited by pretreatment with inhibitors …

Poly(propyleneimine) dendrimer and dendrosome mediated genetic immunization against hepatitis B.

The purpose of the present research work is to explore the potential of dendrosomes in genetic immunization against hepatitis B. Plasmid DNA encoding pRc/CMV-HBs[S] (5.6 kb), encoding the small region of the hepatitis B surface antigen, was complexed with 5th generation poly(propyleneimine) dendrimer (PPI) in different ratios. Transfection of CHO cells revealed that a ratio of 1:50 for pDNA:PPI was optimum for transfection. Results of cytotoxicity studies showed that the toxicity of PPI-DNA complex was significantly (p<0.05) higher for PPI 75 and PPI 100 as compared to the other PPI-DNA comple…

Differential activation of the connexin 43 promoter by dimers of activator protein-1 transcription factors in myometrial cells.

The expression of activator protein-1 (AP-1) transcription factors is increased in the myometrium at term and may therefore regulate the expression of genes, such as connexin 43 (Cx43), required for the onset of labor. The region upstream of the mouse, rat, and human Cx43 genes contains two consensus AP-1 binding sequences, a proximal AP-1, located close to the TATA box, and a distal AP-1, 1 kb upstream. A transient transfection system was developed in which Syrian hamster myometrial cells were transfected with Cx43 promoter-luciferase constructs in combination with expression vectors for the …

Modulating action of the new polymorphism L436F detected in the GLB1 gene of a type-II GM1 gangliosidosis patient.

We report the modulating action of the L436F new polymorphism identified in the GLB1 gene of a patient affected by GM1 gangliosidosis with o … cells, and Western blots. Expression study of the R68W mutated allele resulted in no GLB1 activity. Transfection with a vector carrying the R201C mutation gave rise to a residual GLB1 activity, which, interestingly, was severely reduced in transfection with the L436F/R201C allele. These expression studies, together with co-transfection experiments, suggest tha …

Functional analysis of bone morphogenetic protein type II receptor mutations underlying primary pulmonary hypertension.

A wide range of mutations in the type II receptor for bone morphogenetic protein (BMPR-II) have been shown to underlie primary pulmonary hypertension. To determine the mechanism of altered BMPR-II function, we employed transient transfection studies in cell lines and primary cultures of pulmonary vascular smooth muscle cells using green fluorescent protein (GFP)-tagged wild-type and mutant BMPR2 constructs and confocal microscopy to localize receptors. Substitution of cysteine residues in the ligand binding or kinase domain prevented trafficking of BMPR-II to the cell surface, and reduced bind…

Role of calmodulin in HIV-potentiated Fas-mediated apoptosis.

The recently demonstrated extraordinary rate of turnover of T cells in human immunodeficiency virus (HIV)-1-infected patients and the appare … Fas-mediated apoptosis, the major form of apoptosis in lymphocytes. Our studies have shown that 1) transfection of H9 and MOLT-4 cells with a non-infectious HIV proviral clone, pFN, which expresses wild-type gp160, leads to enhanced Fas-mediated apoptosis, 2) transfection of MOLT-4 cells with a pFN construct pFN delta 147, which expresses a carboxyl-terminall …

Targeted antiangiogenesis gene therapy using targeted cationic microbubbles conjugated with CD105 antibody compared with untargeted cationic and neutral microbubbles.

OBJECTIVE: This study aimed to develop targeted cationic microbubbles conjugated with a CD105 antibody (CMB105) for use in targeted vascular … statin gene was selected for its outstanding antiangiogenesis effect. For in vitro experiments, the transfection efficiency and cell cycle were analyzed using flow cytometry, and the transcription and expression of endostatin were measured by qPCR and Western blotting, respectively. Vascular tube cavity formation and tumor cell invasion were used to evaluate t …

[Research of chromosome aberration in Chinese hamster lung fibroblast induced by VEGF siRNA].

OBJECTIVE: To explore the effects of VEGF siRNA on chromosome aberration in Chinese hamster lung fibroblasts (CHL). METHODS: The chromosome aberrations were observed after CHL were transfected with VEGF siRNA for 24 hours and 48h. RESULTS: There were a questionable positive in 100nmol/L VEGF siRNA after VEGF siRNA were transfected for 24h and the chromosome aberration rate was 6 percentages. Both 50nmol/L VEGF siRNA and 100nmol/L VEGF siRNA caused the questionable positive after VEGF siRNAs were transfected for 48h, and the chromosome aberration rates was 6 percentages and 10 percentages sep

Effect of vector-expressed shRNAs on hTERT expression.

AIM: To study the effect of short hairpin RNAs (shRNAs) expressed from DNA vector on hTERT expression. METHODS: Oligonucleotides coding for four shRNAs against hTERT were cloned into a mammalian shRNA expression vector pUC18U6 to form pUC18U6ht1-4, which were then introduced into HepG2 cells by using liposome-mediated transfection. HepG2 cells transfected by pUC18U6 and pUC18U6GFPsir, which expressed shRNA against green fluorescent protein (GFP), were used as controls. hTERT mRNA in the transfected cells were quantified by using real-time fluorescent RT-PCR. RESULTS: Among the four shRNAs ag…

Induction of artificial cancer stem cells from tongue cancer cells by defined reprogramming factors.

BACKGROUND: The cancer stem cells (CSCs), a small subpopulation of cells in tumor are responsible for the tumor initiation, growth, recurrence and metastasis of cancer, as well as resistance of cancers to drugs or radiotherapy. CSCs are an important target for the development of novel strategies in cancer treatment. However, CSCs-targeted new anti-cancer drug discovery is currently hindered by the lack of easy and reliable methods for isolating, collecting and maintaining sufficient number of CSCs. Here, we examined whether introduction of defined reprogramming factors (Oct4, shp53, Sox2, Klf4

Development and validation of a DNA-launched, intron-stabilized Langat virus infectious clone and reporter viruses.

Langat virus (LGTV) is a tick-borne member of the Flaviviridae family and a biosafety level 2 surrogate for studying tick-borne encephalitis virus (TBEV) replication and pathogenesis. Here, we report the construction of a plasmid encoding a human cytomegalovirus (HCMV) promoter-driven LGTV cDNA that initiates infection following direct transfection of mammalian cells. Incorporation of three introns eliminated viral cDNA-associated toxicity in bacteria, enabling stable propagation of the full-length plasmid. Transfection of this construct resulted in high-level production of infectious LGTV, wh…

Hydroxyapatite nanoparticle mediated delivery of full length dystrophin gene as a potential therapeutic for the treatment of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a severe genetic disorder characterized by progressive muscle degeneration, primarily affecting young m … both mouse myotubes and patient-derived cells, with sustained expression evident up to 5 days post transfection. The patient-derived myotubes also showed dystrophin protein production 7 days post transfection. These findings suggest R-HAp nanoparticles as a promising and cost-effective alternative for DMD treatment, highlighting their potential for overcoming …

Crosslinked self-assemblies of lipoid acid-substituted low molecular weight (1800 Da) polyethylenimine as reductive-sensitive non-viral gene vectors.

In this study, amphiphilic polyethylenimine-graft-thioctic acid (PEI-TA) and polyethylenimine-graft-lauric acid (PEI-LA) were synthesized. Both PEI-TA and PEI-LA could self-assemble into micelles. Due to the existence of disulfide-linked rings at the end of hydrophobic moieties, PEI-TA could form stable micelles with disulfide crosslinked cores (PEI-TA-SS). In comparison with the PEI-LA micelle, PEI-TA-SS possessed higher DNA binding ability according to the gel retardation assay and heparin replacement assay. In vitro transfection experiments indicated that PEI-TA-SS showed comparably high tr…

Heat shock protein 70 (Hsp70)-stimulated deoxycytidine deaminases from a human lymphoma cell but not the activation-induced cytidine deaminase (AID) from Ramos 6.4 human Burkitt's lymphoma cells.

Deoxycytidine deaminase enzyme activity was reduced in lysates of human leukemic THP1 cells 24 h after transfection with siRNA designed to inhibit cell synthesis of heat shock protein 70 (Hsp70)1a and Hsp701b. The cytidine deaminase enzyme activity from the cell lysates was purified from an affinity column which contained bound single-stranded oligodeoxycytidylic acid. Deficient enzyme activity in certain elution fractions from the siRNA-transfected cells was restored by including recombinant HSP 70 in the assays. Enzyme activity in some other fractions was increased after siRNA transfection. …

Sub-genomic replicon and virus-like particles of Omsk hemorrhagic fever virus.

Omsk hemorrhagic fever virus (OHFV) is a member of the tick-borne encephalitis serocomplex of flaviviruses, and causes hemorrhagic disease in humans. To investigate the molecular mechanisms involved in OHFV pathogenesis, we constructed several subgenomic OHFV replicons containing large deletions in the structural region. Replicon RNA was introduced into BHK cells by transfection and the production of viral proteins was monitored by IFA. GFP and luciferase genes were inserted into the OHFV replicon, and these reporter genes were expressed in cells harboring replicating replicon RNA. OHFV replic…

Enhancement of gene transfer to cervical cancer cells using transferrin-conjugated liposome.

Transferrin-conjugated cationic liposome (Tf-DDAB liposome) was developed as a targeted gene delivery system by using heterobifunctional cro … atios, and proved that the 6:1 weight ratio formulation gave the most neutralized complex. In vitro transfection was done in human cervical cancer cells, HeLa, and the transfection efficiency of Tf-liposome was found to be fivefold higher than that of unconjugated (plain) DDAB liposome and twofold higher than that of Lipofectin. In conclusion, a target-oriente …

Rodent complementation group 8 (ERCC8) corresponds to Cockayne syndrome complementation group A.

US31 is a UV-sensitive mutant cell line (rodent complementation group 8) derived from a mouse T cell line L5178Y. We analyzed removal kinetics for UV-induced cyclobutane pyrimidine dimers and (6-4) photoproducts in US31 cells using monoclonal antibodies against these photoproducts. While nearly all (6-4) photoproducts were repaired within 6 h after UV-irradiation, more than 70% of cyclobutane pyrimidine dimers remained unrepaired even 24 h after UV-irradiation. These kinetics resembled those of Cockayne syndrome (CS) cells. Since US31 cells had a low efficiency of cell fusion and transfection,…

Построение олигодендроцитов, подобных прекурсорам, из эмбриональных фибробатластов мыши с использованием miR-7a Имитация.

ЦЕЛИ: Разработка эффективных стратегий повышения эффективности мер по ликвидации последствий по-прежнему является одной из основных терапевтических проблем в связи с такими демиелинизирующими расстройствами, как множественный склероз (МС). МикроРНК (миРНК) стали мощным регулятором клеточной судьбы. Было показано, что miR-7a , в значительной степени выражается в олигодендроцитных клетках-прекурсорах (ОПК). Цель этого исследования заключалась в изучении вопроса о том, является ли завышенное выражение мнений miR-7a :: может перепрограммировать эмбриональные фибробатласты мыши (МЭФ) в похожие на O

Совпадение в аннотации оригинала
OBJECTIVES: Developing effective strategies to enhance remyelination remains a major therapeutic challenge in demyelinating disorders such a … , and transfected with miR-7a mimics using LipofectamineTM 3000 or LipofectamineTM RNAiMAX reagent. Transfection efficiency was assessed by qRT-PCR analysis of miR-7a expression levels. Cells were then cultured in OPC induction medium and assessed for NG2 expression at 14 and 21 days post-transfection. Differentiation potential was examined by transferring ind …

Syncytial therapeutics: Receptor-specific and direct-to-cytosol biologic drug delivery mediated by measles fusion complex.

This work explores cell-cell fusion mediated by measles virus (MeV) as a potential new cell therapy modality that achieves direct-to-cytosol … et" and CHO "target" cells exogenously expressing the measles target SLAM (CHO-SLAM) by mono- or co-transfection of each cell type with plasmids encoding MeV-H and MeV-F. Fusion was observed only in CHO-SLAM cells which were co-transfected with both plasmids, which verified receptor-specificity without false-triggering of fusion in co-transfected "non-target" …