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Does recovery in the spectral characteristics of GdnHCl-denatured Bacillus licheniformis α-amylase due to added calcium point towards protein stabilization?

Halim AA, Feroz SR, Tayyab S

Biosci Biotechnol Biochem, 2013;77(1):87-96.
PMID: 23291750

Treatment of Bacillus licheniformis α-amylase (BLA) with guanidine hydrochloride (GdnHCl) produced both denatured and aggregated forms of the enzyme as studied by circular dichroism, fluorescence, UV difference spectroscopy, size exclusion chromatography (SEC), and enzymatic activity. The presence of CaCl(2) in the incubation mixture produced significant recovery in spectral signals, being complete in presence of 10 mM CaCl(2), as well as in enzymatic activity, which is indicative of protein stabilization. However, the SEC results obtained with GdnHCl-denatured BLA both in the absence and the presence of 10 mM CaCl(2) suggested significant aggregation of the protein in the absence of CaCl(2) and disaggregation in its presence. Although partial structural stabilization with significant retention of enzymatic activity was observed in the presence of calcium, it was far from the native state, as reflected by spectral probes. Hence, spectral results as to BLA stabilization should be treated with caution in the presence of aggregation.

Matched MeSH terms: Spectrometry, Fluorescence
A variant e13a3 BCR-ABL1 fusion transcript in refractory adult B-cell acute lymphoblastic leukemia achieving complete remission with CAR-Tcell therapy

Phan CL, Tan SN, Tan SM, Kadir SSSA, Ramli NLM, Lim TO, et al.

Cancer Genet, 2021 01;250-251:20-24.
PMID: 33220656 DOI: 10.1016/j.cancergen.2020.11.003

Acute lymphoblastic leukemia (ALL) cases with e13a3 fusion transcripts are extremely rare. We report a 24-year-old male with Ph-positive (Ph+) ALL with an aberrant e13a3 fusion transcript treated with CD19-specific chimeric antigen receptor T-cell (CAR-T) therapy. He developed refractory disease post-chemotherapy induction, andreceived allogeneic hematopoietic stem cell transplantation (allo-HSCT) after salvage with imatinib in combination with chemotherapy regimen. Unfortunately, the patient relapsed after +90 days post-transplant. He was consented to CAR-T therapy trial and achieved complete remission, highlighting the efficacy of CAR-T treatment in relapsed-refractory B-ALL irrespective of the underlying genetic drivers in leukemia cells .

Matched MeSH terms: In Situ Hybridization, Fluorescence
Quantification of (+)-calanolide A, a novel and naturally occurring anti-HIV agent, by high-performance liquid chromatography in plasma from rat, dog and human

Xu ZQ, Norris KJ, Weinberg DS, Kardatzke J, Wertz P, Frank P, et al.

J Chromatogr B Biomed Sci Appl, 2000 Jun 09;742(2):267-75.
PMID: 10901131

A HPLC method was validated for quantification of (+)-calanolide A (1), a novel anti-HIV agent, in rat, dog and human plasma. The synthetic intermediate (+/-)-12-oxocalanolide A (2) was found to be a suitable internal standard. Compounds were extracted from plasma using a solid-phase C(18) cartridge and quantified over the assay range of 12.5 to 800 ng/ml. The method was utilized to determine (+)-calanolide A pharmacokinetics in rats, dogs and humans. This is the first report of a validated HPLC assay for determination of (+)-calanolide A concentrations in rat and dog plasma as well as human plasma obtained from clinical trials. There was no evidence of in vivo epimerization of (+)-calanolide A to its inactive epimer (+)-calanolide B (3).

Matched MeSH terms: Spectrometry, Fluorescence
Effects of testosterone on mean arterial pressure and aquaporin (AQP)-1, 2, 3, 4, 6 and 7 expressions in the kidney of orchidectomized, adult male Sprague-Dawley rats

Loh SY, Giribabu N, Gholami K, Salleh N

Arch Biochem Biophys, 2017 Jan 15;614:41-49.
PMID: 28024836 DOI: 10.1016/j.abb.2016.12.008

We hypothesized that higher blood pressure in males than females could be due to testosterone effects on aquaporin (AQP) expression in kidneys.
METHODS: Orchidectomized adult male Sprague-Dawley (SD) rats received seven days subcutaneous testosterone treatment (125 μg/kg/day or 250 μg/kg/day), with or without flutamide or finasteride. Following completion of treatment, MAP was determined in rats under anaesthesia via carotid artery cannulation. In another cohort of rats, kidneys were removed following sacrifice and AQP-1, 2, 3, 4, 6 and 7 protein and mRNA levels were determined by Western blotting and Real-time PCR respectively. Distribution of AQP subunits' protein in the nephrons were visualized by immunofluorescence.
RESULTS: Testosterone caused MAP, AQP-1, 2, 4, 6 and 7 protein and mRNA levels in kidneys to increase while AQP-3 protein and mRNA levels in kidneys to decrease (p

Matched MeSH terms: Microscopy, Fluorescence
An Overview of Circular RNAs

Awasthi R, Singh AK, Mishra G, Maurya A, Chellappan DK, Gupta G, et al.

Adv Exp Med Biol, 2018 9 28;1087:3-14.
PMID: 30259353 DOI: 10.1007/978-981-13-1426-1_1

Circular RNAs (cirRNAs) are long, noncoding endogenous RNA molecules and covalently closed continuous loop without 5'-3' polarity and polyadenylated tail which are largely concentrated in the nucleus. CirRNA regulates gene expression by modulating microRNAs and functions as potential biomarker. CirRNAs can translate in vivo to link between their expression and disease. They are resistant to RNA exonuclease and can convert to the linear RNA by microRNA which can then act as competitor to endogenous RNA. This chapter summarizes the evolutionary conservation and expression of cirRNAs, their identification, highlighting various computational approaches on cirRNA, and translation with a focus on the breakthroughs and the challenges in this new field.

Matched MeSH terms: In Situ Hybridization, Fluorescence
Macrophage response and surface analysis of dental cementum after treatment with high intensity focused ultrasound

Daood U, Fawzy AS

Arch Oral Biol, 2019 Feb;98:195-203.
PMID: 30502562 DOI: 10.1016/j.archoralbio.2018.10.019

OBJECTIVE: To investigate effects of HIFU on macrophage phenotype, surface micro-topography and nano-scale surface mechanical properties of dental cementum.
MATERIALS AND METHODS: Root discs (2 mm thickness) were cut apical to CEJ and sectioned into quadrants. HIFU setup with bowl-shaped piezo ceramic transducer submerged in a water tank was used for exposure on each specimen for 15 s, 30 s or 60 s. The specimens of the control group were left without any HIFU exposure. HIFU was generated with a continuous sinusoidal wave of 120Vpp amplitude, 250 KHZ resonance-frequency and highest ultrasonic pressure of ∼10 bar at the focus. Specimens for SEM were viewed, and micro-topography characterization performed, using AFM and Ra parameter and surface area (SA) calculated by specialized SPM surface analysis software. For nano-indentation testing, experiments were carried out using AFM. Macrophage cell isolation and culturing was performed on cementum to receive the HIFU treatment at different time periods. Raman spectroscopy were scanned to create spectra perpendicular to the cementum substrate to analyze generation of standard spectra for Raman intensity ratio of hydroxyapatite normalized to the peaks ν1 960 cm-1. Data was expressed as means ± standard deviations and analyzed by one-way ANOVA in term of Ra, SA, H and Er. Different points for fluorescence intensity ratio were analyzed by Raman using Wilcoxon rank sum test.
RESULTS: HIFU exposure at 60 s removed the smear layer and most of cementum appeared smoothened. AFM characterisation, showed a slight decrease in the irregularity of the surface as exposure time increased. Intact macrophages can be identified in control and all experimental HIFU groups. The level of fluorescence for the control and HIFU 15 and 30 s were low as compared to HIFU 60 s.
CONCLUSION: If HIFU can be successfully implemented, it may be a possible alternative to current methods used in periodontal therapy to achieve smooth root surfaces.

Matched MeSH terms: Fluorescence
Fulltext Comprehensive Genetic Results for Primary Immunodeficiency Disorders in a Highly Consanguineous Population

Al-Herz W, Chou J, Delmonte OM, Massaad MJ, Bainter W, Castagnoli R, et al.

Front Immunol, 2018;9:3146.
PMID: 30697212 DOI: 10.3389/fimmu.2018.03146

Objective: To present the genetic causes of patients with primary immune deficiencies (PIDs) in Kuwait between 2004 and 2017. Methods: The data was obtained from the Kuwait National Primary Immunodeficiency Disorders Registry. Genomic DNA from patients with clinical and immunological features of PID was sequenced using Sanger sequencing (SS), next generation sequencing (NGS) of targeted genes, whole exome sequencing (WES), and/or whole genome sequencing (WGS). Functional assays were utilized to assess the biologic effect of identified variants. Fluorescence in situ hybridization (FISH) for 22q11.2 deletion and genomic hybridizations arrays were performed when thymic defects were suspected. Results: A total of 264 patients were registered during the study period with predominance of patients with immunodeficiencies affecting cellular and humoral immunity (35.2%), followed by combined immunodeficiencies with associated syndromic features (24%). Parental consanguinity and family history suggestive of PID were reported in 213 (81%) and 145 patients (55%), respectively. Genetic testing of 206 patients resulted in a diagnostic yield of 70%. Mutations were identified in 46 different genes and more than 90% of the reported genetic defects were transmitted by in an autosomal recessive pattern. The majority of the mutations were missense mutations (57%) followed by deletions and frame shift mutations. Five novel disease-causing genes were discovered. Conclusions: Genetic testing should be an integral part in the management of primary immunodeficiency patients. This will help the delivery of precision medicine and facilitate proper genetic counseling. Studying inbred populations using sophisticated diagnostic methods can allow better understanding of the genetics of primary immunodeficiency disorders.

Matched MeSH terms: In Situ Hybridization, Fluorescence
Fulltext Diversity, enumeration, and isolation of Arcobacter spp. in the giant abalone, Haliotis gigantea

Mizutani Y, Iehata S, Mori T, Oh R, Fukuzaki S, Tanaka R

Microbiologyopen, 2019 10;8(10):e890.
PMID: 31168933 DOI: 10.1002/mbo3.890

Arcobacter have been frequently detected in and isolated from bivalves, but there is very little information on the genus Arcobacter in the abalone, an important fishery resource. This study aimed to investigate the genetic diversity and abundance of bacteria from the genus Arcobacter in the Japanese giant abalone, Haliotis gigantea, using molecular methods such as Arcobacter-specific clone libraries and fluorescence in situ hybridization (FISH). Furthermore, we attempted to isolate the Arcobacter species detected. Twelve genotypes of clones were obtained from Arcobacter-specific clone libraries. These sequences are not classified with any other known Arcobacter species including pathogenic Arcobacter spp., A. butzleri, A. skirrowii, and A. cryaerophilus, commonly isolated or detected from bivalves. From the FISH analysis, we observed that ARC94F-positive cells, presumed to be Arcobacter, accounted for 6.96 ± 0.72% of all EUB338-positive cells. In the culture method, three genotypes of Arcobacter were isolated from abalones. One genotype had a similarity of 99.2%-100.0% to the 16S rRNA gene of Arcobacter marinus, while the others showed only 93.3%-94.3% similarity to other Arcobacter species. These data indicate that abalones carry Arcobacter as a common bacterial genus which includes uncultured species.

Matched MeSH terms: In Situ Hybridization, Fluorescence
In Vitro Investigation of Influences of Chitosan Nanoparticles on Fluorescein Permeation into Alveolar Macrophages

Chachuli SH, Nawaz A, Shah K, Naharudin I, Wong TW

Pharm Res, 2016 06;33(6):1497-508.
PMID: 26951565 DOI: 10.1007/s11095-016-1893-5

PURPOSE: Pulmonary infection namely tuberculosis is characterized by alveolar macrophages harboring a large microbe population. The chitosan nanoparticles exhibit fast extracellular drug release in aqueous biological milieu. This study investigated the matrix effects of chitosan nanoparticles on extracellular drug diffusion into macrophages.
METHODS: Oligo, low, medium and high molecular weight chitosan nanoparticles were prepared by nanospray drying technique. These nanoparticles were incubated with alveolar macrophages in vitro and had model drug sodium fluorescein added into the same cell culture. The diffusion characteristics of sodium fluorescein and nanoparticle behavior were investigated using fluorescence microscopy, scanning electron microscopy, differential scanning calorimetry and Fourier transform infrared spectroscopy techniques.
RESULTS: The oligochitosan nanoparticles enabled macrophage membrane fluidization with the extent of sodium fluorescein entry into macrophages being directly governed by the nanoparticle loading. Using nanoparticles made of higher molecular weight chitosan, sodium fluorescein permeation into macrophages was delayed due to viscous chitosan diffusion barrier at membrane boundary.
CONCLUSION: Macrophage-chitosan nanoparticle interaction at membrane interface dictates drug migration into cellular domains.

Matched MeSH terms: Microscopy, Fluorescence
Ratiometric Detection of microRNA Using Hybridization Chain Reaction and Fluorogenic Silver Nanoclusters

Wong ZW, Ng JF, New SY

Chem Asian J, 2021 Dec 13;16(24):4081-4086.
PMID: 34668337 DOI: 10.1002/asia.202101145

miRNA (miR)-155 is a potential biomarker for breast cancers. We aimed at developing a nanosensor for miR-155 detection by integrating hybridization chain reaction (HCR) and silver nanoclusters (AgNCs). HCR serves as an enzyme-free and isothermal amplification method, whereas AgNCs provide a built-in fluorogenic detection probe that could simplify the downstream analysis. The two components were integrated by adding a nucleation sequence of AgNCs to the hairpin of HCR. The working principle was based on the influence of microenvironment towards the hosted AgNCs, whereby unfolding of hairpin upon HCR has manipulated the distance between the hosted AgNCs and cytosine-rich toehold region of hairpin. As such, the dominant emission of AgNCs changed from red to yellow in the absence and presence of miR-155, enabling a ratiometric measurement of miR with high sensitivity. The limit of detection (LOD) of our HCR-AgNCs nanosensor is 1.13 fM in buffered solution. We have also tested the assay in diluted serum samples, with comparable LOD of 1.58 fM obtained. This shows the great promise of our HCR-AgNCs nanosensor for clinical application.

Matched MeSH terms: Spectrometry, Fluorescence
Synthesis, Molecular Docking and Biological Activity Evaluation of Alkoxy Substituted Chalcone Derivatives: Potential Apoptosis Inducing Agent on MCF-7 Cells

Khairul WM, Hashim F, Mohammed M, Shah NSMN, Johari SATT, Rahamathullah R, et al.

Anticancer Agents Med Chem, 2021;21(13):1738-1750.
PMID: 33176667 DOI: 10.2174/1871520620999201110190709

INTRODUCTION: In this contribution, a series of alkoxy substituted chalcones were successfully designed, synthesized, spectroscopically characterized and evaluated for their cytotoxicity potential in inhibiting the growth of MCF-7 cells.
OBJECTIVE: In order to investigate the influence between electron density in conjugated π-systems and biological activities, different withdrawing substituents, namely Nitro (NO2), Cyano (C≡N) and trifluoromethyl (CF3) were introduced in the chalcone-based molecular system.
METHODS: All the derivatives were then tested on MCF-7 cell line using the fluorescence microscopy-based cytotoxicity analyses.
RESULTS: The preliminary findings showed that both -NO2 and -CF3 substituents revealed their potential to inhibit the growth of MCF-7 with IC;50 values of 14.75 and 13.75 μg/ml, respectively. In addition, the morphological changes of MCF-7 cells were observed in response to alkoxy substituted chalcone treatment through an induction of apoptosis pathway with cell blebbing, phosphatidylserine exposure and autophagic activity with acidification of lysosomal structure. Intermolecular interaction based on in silico investigation on nitro, trifluoromethyl and cyano based chalcones exhibited several types of interactions with tumor necrosis factor receptor (PDB: 1EXT) protein and high hydrogen bond in the molecule-receptor interaction have given significant impact towards their toxicity on MCF-7 cells.
CONCLUSION: Significantly, these types of chalcones exhibited ideal and high potential to be further developed as anti-cancer agents.

Matched MeSH terms: Microscopy, Fluorescence
Cell Wall-Treated Lactococcus lactis Increases the Plasmid Transfer Efficiency of Internal Ribosome Entry Site-Incorporated Lactococcal Bicistronic Vector into DF1 Cells

Mat Isa N, Abdul Mutalib NE, Alitheen NB, Song AA, Rahim RA

J. Mol. Microbiol. Biotechnol., 2017;27(4):246-251.
PMID: 29055951 DOI: 10.1159/000481257

This study demonstrates that cell wall treatment of Lactococcus lactis harbouring the internal ribosome entry site-incorporated lactococcal bicistronic vector pNZ:VIG mediated the delivery of genes into an eukaryotic cell line, DF1 cells, through bactofection. Bactofection analysis showed that the pNZ:VIG plasmid in L. lactis can be transferred into DF1 cells and that both the VP2 and gfp genes cloned in the plasmid can be transcribed and translated. The protein band relative to the Mr of VP2 protein (49 kDa) was successfully detected via Western blot analysis, while green fluorescence was successfully detected using a fluorescence microscope. The intensity of the bands detected increased for samples treated with both 1.5% (w/v) glycine and 10 μg/mL of lysozyme when compared to L. lactis treated with glycine alone and without treatment. Cell wall treatment of L. lactis with a combination of both glycine and lysozyme was not only shown to mediate plasmid transfer to DF1 cells, but also to increase the plasmid transfer efficiency.

Matched MeSH terms: Microscopy, Fluorescence
Biophysical and computational characterization of vandetanib-lysozyme interaction

Kabir MZ, Hamzah NAB, Ghani H, Mohamad SB, Alias Z, Tayyab S

Spectrochim Acta A Mol Biomol Spectrosc, 2018 Jan 15;189:485-494.
PMID: 28843881 DOI: 10.1016/j.saa.2017.08.051

Interaction of an anticancer drug, vandetanib (VDB) with a ligand transporter, lysozyme (LYZ) was explored using multispectroscopic techniques, such as fluorescence, absorption and circular dichroism along with computational analysis. Fluorescence data and absorption results confirmed VDB-LYZ complexation. VDB-induced quenching was characterized as static quenching based on inverse correlation of KSV with temperature as well as kq values. The complex was characterized by the weak binding constant (Ka=4.96-3.14×103M-1). Thermodynamic data (ΔS=+12.82Jmol-1K-1; ΔH=-16.73kJmol-1) of VDB-LYZ interaction revealed participation of hydrophobic and van der Waals forces along with hydrogen bonds in VDB-LYZ complexation. Microenvironmental perturbations around tryptophan and tyrosine residues as well as secondary and tertiary structural alterations in LYZ upon addition of VDB were evident from the 3-D fluorescence, far- and near-UV CD spectral analyses, respectively. Interestingly, addition of VDB to LYZ significantly increased protein's thermostability. Molecular docking results suggested the location of VDB binding site near the LYZ active site while molecular dynamics simulation results suggested stability of VDB-LYZ complex. Presence of Mg2+, Ba2+ and Zn2+ was found to interfere with VDB-LYZ interaction.

Matched MeSH terms: Spectrometry, Fluorescence
Fulltext Evolutionary and functional analysis of RBMY1 gene copy number variation on the human Y chromosome

Shi W, Louzada S, Grigorova M, Massaia A, Arciero E, Kibena L, et al.

Hum Mol Genet, 2019 Aug 15;28(16):2785-2798.
PMID: 31108506 DOI: 10.1093/hmg/ddz101

Human RBMY1 genes are located in four variable-sized clusters on the Y chromosome, expressed in male germ cells and possibly associated with sperm motility. We have re-investigated the mutational background and evolutionary history of the RBMY1 copy number distribution in worldwide samples and its relevance to sperm parameters in an Estonian cohort of idiopathic male factor infertility subjects. We estimated approximate RBMY1 copy numbers in 1218 1000 Genomes Project phase 3 males from sequencing read-depth, then chose 14 for valid ation by multicolour fibre-FISH. These fibre-FISH samples provided accurate calibration standards for the entire panel and led to detailed insights into population variation and mutational mechanisms. RBMY1 copy number worldwide ranged from 3 to 13 with a mode of 8. The two larger proximal clusters were the most variable, and additional duplications, deletions and inversions were detected. Placing the copy number estimates onto the published Y-SNP-based phylogeny of the same samples suggested a minimum of 562 mutational changes, translating to a mutation rate of 2.20 × 10-3 (95% CI 1.94 × 10-3 to 2.48 × 10-3) per father-to-son Y-transmission, higher than many short tandem repeat (Y-STRs), and showed no evidence for selection for increased or decreased copy number, but possible copy number stabilizing selection. An analysis of RBMY1 copy numbers among 376 infertility subjects failed to replicate a previously reported association with sperm motility and showed no significant effect on sperm count and concentration, serum follicle stimulating hormone (FSH), luteinizing hormone (LH) and testosterone levels or testicular and semen volume. These results provide the first in-depth insights into the structural rearrangements underlying RBMY1 copy number variation across diverse human lineages.

Matched MeSH terms: In Situ Hybridization, Fluorescence
18q21 Rearrangement and trisomy 3 in extranodal B-cell lymphomas: a study using a fluorescent in situ hybridisation technique

Tai YC, Tan JA, Peh SC

Virchows Arch, 2004 Nov;445(5):506-14.
PMID: 15365830

t(11;18)(q21;q21) Translocation and trisomy 3 are the most common chromosomal aberrations reported in low-grade mucosa-associated lymphoid tissue (MALT) lymphoma. The current study aims to investigate the frequency of these chromosomal aberrations in a series of 52 extranodal B-cell lymphomas. The tumours were categorised into three histological grades: grade 1 (low-grade lymphoma of MALT type), grade 2 [diffuse large B-cell lymphoma (DLBCL) with MALT component] and grade 3 (DLBCL without MALT component). Fluorescence in situ hybridisation analyses on paraffin tissue sections were performed using a locus-specific probe for the 18q21 region and a centromeric probe for chromosome 3. The 18q21 rearrangement was detected in 9 of 40 (23%) cases, including 7 of 23 (30%) grade-1 and 2 of 11 (18%) grade-3 tumours. Amplification of the 18q21 region was detected in 10 of 40 (25%) cases, and trisomy 3 was detected in 9 of 34 (26%) cases. Amplification of the 18q21 region may be an important alternative pathogenetic pathway in MALT lymphoma and was found almost exclusively in tumours without 18q21 rearrangement. Our study showed that tumours with 18q21 rearrangement and 18q21 amplification develop along two distinct pathways, and the latter was more likely to transform into high-grade tumours upon acquisition of additional genetic alterations, such as trisomy 3. Trisomy 3 was more frequently found in coexistence with 18q21 abnormalities, suggesting that it was more likely to be a secondary aberration.

Matched MeSH terms: In Situ Hybridization, Fluorescence
Fulltext Effect of Root Restriction on the Growth, Photosynthesis Rate, and Source and Sink Relationship of Chilli (Capsicum annuum L.) Grown in Soilless Culture

Zakaria NI, Ismail MR, Awang Y, Megat Wahab PE, Berahim Z

Biomed Res Int, 2020;2020:2706937.
PMID: 32090071 DOI: 10.1155/2020/2706937

Chilli (Capsicum annum L.) plant is a high economic value vegetable in Malaysia, cultivated in soilless culture containers. In soilless culture, the adoption of small container sizes to optimize the volume of the growing substrate could potentially reduce the production cost, but will lead to a reduction of plant growth and yield. By understanding the physiological mechanism of the growth reduction, several potential measures could be adopted to improve yield under restricted root conditions. The mechanism of growth reduction of plants subjected to root restriction remains unclear. This study was conducted to determine the physiological mechanism of growth reduction of root-restricted chilli plants grown in polyvinyl-chloride (PVC) column of two different volumes, 2392 cm3(root-restricted) and 9570 cm3(control) in soilless culture. Root restriction affected plant growth, physiological process, and yield of chilli plants. Root restriction reduced the photosynthesis rate and photochemical activity of PSII, and increased relative chlorophyll content. Limited root growth in root restriction caused an accumulation of high levels of sucrose in the stem and suggested a transition of the stem as a major sink organ for photoassimilate. Growth reduction in root restriction was not related to limited carbohydrate production, but due to the low sink demand from the roots. Reduction of the total yield per plant about, 23% in root restriction was concomitant, with a slightly increased harvest index which reflected an increased photoassimilate partitioning to the fruit production and suggested more efficient fruits production in the given small plant size of root restriction.

Matched MeSH terms: Fluorescence
Fulltext Gene transfer into the lung by nanoparticle dextran-spermine/plasmid DNA complexes

Abdullah S, Wendy-Yeo WY, Hosseinkhani H, Hosseinkhani M, Masrawa E, Ramasamy R, et al.

J Biomed Biotechnol, 2010;2010:284840.
PMID: 20617146 DOI: 10.1155/2010/284840

A novel cationic polymer, dextran-spermine (D-SPM), has been found to mediate gene expression in a wide variety of cell lines and in vivo through systemic delivery. Here, we extended the observations by determining the optimal conditions for gene expression of D-SPM/plasmid DNA (D-SPM/pDNA) in cell lines and in the lungs of BALB/c mice via instillation delivery. In vitro studies showed that D-SPM could partially protect pDNA from degradation by nuclease and exhibited optimal gene transfer efficiency at D-SPM to pDNA weight-mixing ratio of 12. In the lungs of mice, the levels of gene expression generated by D-SPM/pDNA are highly dependent on the weight-mixing ratio of D-SPM to pDNA, amount of pDNA in the complex, and the assay time postdelivery. Readministration of the complex at day 1 following the first dosing showed no significant effect on the retention and duration of gene expression. The study also showed that there was a clear trend of increasing size of the complexes as the amount of pDNA was increased, where the sizes of the D-SPM/pDNA complexes were within the nanometer range.

Matched MeSH terms: Microscopy, Fluorescence
Fulltext Nipah Virus Matrix Protein Influences Fusogenicity and Is Essential for Particle Infectivity and Stability

Dietzel E, Kolesnikova L, Sawatsky B, Heiner A, Weis M, Kobinger GP, et al.

J Virol, 2016 Mar;90(5):2514-22.
PMID: 26676785 DOI: 10.1128/JVI.02920-15

Nipah virus (NiV) causes fatal encephalitic infections in humans. To characterize the role of the matrix (M) protein in the viral life cycle, we generated a reverse genetics system based on NiV strain Malaysia. Using an enhanced green fluorescent protein (eGFP)-expressing M protein-deleted NiV, we observed a slightly increased cell-cell fusion, slow replication kinetics, and significantly reduced peak titers compared to the parental virus. While increased amounts of viral proteins were found in the supernatant of cells infected with M-deleted NiV, the infectivity-to-particle ratio was more than 100-fold reduced, and the particles were less thermostable and of more irregular morphology. Taken together, our data demonstrate that the M protein is not absolutely required for the production of cell-free NiV but is necessary for proper assembly and release of stable infectious NiV particles.

Matched MeSH terms: Microscopy, Fluorescence
Synthesis, structure, DNA/BSA interaction and in vitro cytotoxic activity of nickel(II) complexes derived from S-allyldithiocarbazate

Nanjundan N, Selvakumar P, Narayanasamy R, Haque RA, Velmurugan K, Nandhakumar R, et al.

J. Photochem. Photobiol. B, Biol., 2014 Dec;141:176-85.
PMID: 25463665 DOI: 10.1016/j.jphotobiol.2014.10.009

Two nickel(II) complexes with formula NiL1 and NiL2 (HL1 = S-allyl-4-methoxybenzylidene hydrazinecarbodithioate, HL2 = S-allyl-1-napthylidenehydrazinecarbodithioate) have been synthesized and characterized by elemental analysis, FT-IR, NMR, UV-vis spectroscopy and ESI mass spectrometry. The crystal structure of complex 1 has been determined by single crystal X-ray diffractometry. Both HL1 and HL2 ligands are coordinated to the metal in thiolate form. In complexes, squareplanar geometry of the nickel is coordinated with two bidentate ligand units acting through azomethine nitrogen and thiolato sulfur atoms. To explore the potential medicinal value of the complexes with calf thymus DNA and bovine serum albumin (BSA) were studied at normal physiological conditions using fluorescence spectral techniques. The DNA binding constant values of the complexes were found in the range from 5.02 × 10(4), 3.54 × 10(4), and the binding affinities are in the following order 1 > 2. In addition, nickel complexes 1 and 2 shows better binding propensity to the bovine serum albumin (BSA) protein, giving a Ksv value 5.8 × 10(4), 4.47 × 10(4) respectively. From the oxidative cleavage of the complexes with pBR322 DNA, it is inferred that the effects of cleavage are dose-dependent. In addition, in vitro cytotoxicity of the complexes assayed against Vero and HeLa cell lines have shown higher cytotoxic activity with the lower IC50 values indicating their efficiency in killing cancer cells even at various concentrations.

Matched MeSH terms: Spectrometry, Fluorescence
Fulltext Cytotoxic constituents from the rhizomes of Curcuma zedoaria

Ahmed Hamdi OA, Syed Abdul Rahman SN, Awang K, Abdul Wahab N, Looi CY, Thomas NF, et al.

ScientificWorldJournal, 2014;2014:321943.
PMID: 25126594 DOI: 10.1155/2014/321943

Curcuma zedoaria also known as Temu putih is traditionally used in food preparations and treatment of various ailments including cancer. The cytotoxic activity of hexane, dichloromethane, ethyl acetate, methanol, and the methanol-soxhlet extracts of Curcuma zedoaria rhizomes was tested on two human cancer cell lines (Ca Ski and MCF-7) and a noncancer cell line (HUVEC) using MTT assay. Investigation on the chemical components in the hexane and dichloromethane fractions gave 19 compounds, namely, labda-8(17),12 diene-15,16 dial (1), dehydrocurdione (2), curcumenone (3), comosone II (4), curcumenol (5), procurcumenol (6), germacrone (7), zerumbone epoxide (8), zederone (9), 9-isopropylidene-2,6-dimethyl-11-oxatricyclo[6.2.1.0(1,5)]undec-6-en-8-ol (10), furanodiene (11), germacrone-4,5-epoxide (12), calcaratarin A (13), isoprocurcumenol (14), germacrone-1,10-epoxide (15), zerumin A (16), curcumanolide A (17), curcuzedoalide (18), and gweicurculactone (19). Compounds (1-19) were evaluated for their antiproliferative effect using MTT assay against four cancer cell lines (Ca Ski, MCF-7, PC-3, and HT-29). Curcumenone (3) and curcumenol (5) displayed strong antiproliferative activity (IC50 = 8.3 ± 1.0 and 9.3 ± 0.3 μg/mL, resp.) and were found to induce apoptotic cell death on MCF-7 cells using phase contrast and Hoechst 33342/PI double-staining assay. Thus, the present study provides basis for the ethnomedical application of Curcuma zedoaria in the treatment of breast cancer.

Matched MeSH terms: Microscopy, Fluorescence

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