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Detection of Apoptotic Circulating Tumor Cells Using in vivo Fluorescence Flow Cytometry
摘要： Most cancer patients die from metastatic disease as a result of a circulating tumor cell (CTC) spreading from a primary tumor through the blood circulation to distant organs. Many studies have demonstrated the tremendous potential of using CTC counts as prognostic markers of metastatic development and therapeutic efficacy. However, it is only the viable CTCs capable of surviving in the blood circulation that can create distant metastasis. To date, little progress has been made in understanding what proportion of CTCs is viable and what proportion is in an apoptotic state. Here, we introduce a novel approach toward in situ characterization of CTC apoptosis status using a multi-color in vivo flow cytometry platform with fluorescent detection for the real-time identification and enumeration of such cells directly in blood flow. The proof of concept was demonstrated with two-color fluorescence flow cytometry (FFC) using breast cancer cells MDA-MB-231 expressing green fluorescein protein (GFP), staurosporine as an activator of apoptosis, Annexin-V apoptotic kit with orange dye color, and a mouse model. The future application of this new platform for real-time monitoring of antitumor drug efficiency is discussed.
关键词： fluorescence flow cytometry,circulating tumor cells,in vivo detection of circulating tumor cells,apoptosis
更新于2025-09-10 09:29:36
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Novel Protective Effects of Cistanche Tubulosa Extract Against Low-Luminance Blue Light-Induced Degenerative Retinopathy
摘要： Background/Aims: Blue light-emitting diode light (BLL)-induced phototoxicity plays an important role in ocular diseases and causes retinal degeneration and apoptosis in human retinal pigment epithelial (RPE) cells. Cistanche tubulosa extract (CTE) is a traditional Chinese medicine with many beneficial protective properties; however, few studies have examined the ocular protective roles of CTE. In this study, we investigated the mechanisms underlying the effects of CTE on BLL-induced apoptosis in vitro and in vivo. Methods: RPE cells were applied in the current in vitro study and cell viability was determined by an 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Apoptosis-related protein expression was determined by western blot analysis and immunofluorescence staining. Brown Norway rats were used to examine exposure to commercially available BLL in vivo. Hematoxylin and eosin staining, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and western blot assays were used to examine retinal morphological deformation. Results: CTE significantly inhibited hydrogen peroxide-, tert-butyl hydroperoxide-, sodium azide-, and BLL-induced RPE damage. Further, CTE reduced the expression of apoptotic markers such as cleaved caspase-3 and TUNEL staining after BLL exposure by inactivating apoptotic pathways, as shown via immunofluorescent staining. In addition, CTE inhibited the BLL-induced phosphorylation of c-Jun N-terminal kinase, extra signal-related kinases 1/2, and p38 in RPE cells. In vivo, the oral administration of CTE rescued 60-day periodic BLL exposure-induced decrements in retinal thickness and reduced the number of TUNEL-positive cells in the brown Norway rat model. Conclusion: CTE is a potential prophylactic agent against BLL-induced phototoxicity.
关键词： Cistanche tubulosa extract,Blue light,Retinal pigment epithelial cells,Apoptosis
更新于2025-09-10 09:29:36
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Biosynthesis and characterization of copper oxide nanoparticles from indigenous fungi and its effect of photothermolysis on human lung carcinoma
摘要： In this report, copper oxide nanoparticles (TA-CuO NPs) were synthesized using cell-free extract of Trichoderma asperellum and assessed their photothermal induced anticancerous activity. The fungal mediated TA-CuO NPs was confirmed by the surface plasmon resonance at 285-295 nm. The amide (C=O) and aromatic (C=C) groups in secondary metabolites of the extract was found to be an encapsulating or reducing agents for TA-CuO NPs, as indicated by IR spectra. Crystalline nature by cubic face-centered structure of the TA-CuO NPs was confirmed by XRD and their size ranges from 10 to 190 nm and an average of 110 nm by particle size analyzer (PSA). The Ultra HRSEM study revealed spherical shaped TA-CuO NPs. The FETEM results were also in strong agreement with PSA and UHR SEM. The survey-scan spectrum of XPS indicated the presence of C1s (47.83%), Cu2p (16.11%), Na1s (2.2%) and O1s (33.86%). The cell death was significantly found higher in photothermal induced by near-infrared laser (TA-CuO NPs-NIR) treated than that of TA-CuO NPs treatment. The level of ROS (35.62 %) was higher in the treated cells than that of the untreated control, in accordance with the nucleus damage and losses in the mitochondrial membrane potential (ΔΨm). the upregulation of Bcl-2 in untreated cells and Cas-3 in TA-CuO NPs-NIR treated cells was confirmed by western blot analysis. This work agreed the potential biogenic TA-CuO NPs for promising in vitro photothermolysis of cancer cells, for the development of anticancer nanotherapeutics.
关键词： photothermal activity,Trichoderma,copper oxide nanoparticles,human lung carcinoma,apoptosis,anticancer
更新于2025-09-10 09:29:36
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Silica Nanoparticles with Up-conversion Fluorescence Based on Triplet-Triplet Annihilation Mechanism for Specific Recognition of Apoptosis Cells
摘要： Discriminating of live and apoptotic cells is a crucial task in the research of pharmacology, biology, pathology, and medicine science. Recently, up conversion (UC) luminescent materials have appealed much attention due to their unique ability to convert low energy excitation photons to high energy ones. However, UC fluorescence has not been employed in the field of discriminating of live and apoptotic cells. We presented a facile, and cost-less St?ber method to fabricate robust silica nanoparticles (SiO2 UCNPs) exhibiting several merits, such as narrow size distribution, and UC luminescence. SiO2 UCNPs could discriminate live and apoptosis cells by taking advantage of the unique surface property of SiO2 UCNPs for the first time. This work is also the first demonstration of the use of single photon excited UC fluorescence derived from nanoparticles for biological recognition of a specific type of cells.
关键词： Apoptosis Cells,Triplet-Triplet Annihilation Mechanism,Silica Nanoparticles,Up-conversion Fluorescence
更新于2025-09-09 09:28:46
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[IEEE 2018 9th International Conference on Ultrawideband and Ultrashort Impulse Signals (UWBUSIS) - Odessa, Ukraine (2018.9.4-2018.9.7)] 2018 9th International Conference on Ultrawideband and Ultrashort Impulse Signals (UWBUSIS) - The Application of Pulsed Electric Fields and Other Types of Electromagnetic Radiation in Therapy of Cancer
摘要： since the beginning of 21-th century the investigations in the area of cancer therapy by electric fields become more numerous. These investigations may be divided in three groups: studies carried out on cell cultures, studies on animal models and case reports describing applications of pulsed electric fields for treatment of cancer patients. Very important is to chose adequate methods of measuring the cell damage and speaking more generally – cell response to stress. In Kharkov University various investigations in the area of cell response to electromagnetic influences were carried out and some new methods were proposed. In this paper the analysis of recent works in this area is done and perspectives of the research are discussed.
关键词： electromagnetic pulse,cell proliferation,cell membrane permeability,cell response to electromagnetic stress,apoptosis,cancer therapy,necrosis
更新于2025-09-09 09:28:46
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Flow Cytometry - Select Topics || Sperm Flow Cytometry: Beyond Human Fertilization and Embryo Development
摘要： Male infertily is a contributing factor in up to 50% of all infertility cases, a solo cause in about 30% of them. Therefore, new and improved diagnostic methods that reduce operator variability regarding sperm defects that are not accesible by the conventional microscope scoring should be evaluated. Assisted reproductive technology (ART) has been involved in the description of alternative pathways in basic cellular functions. it is important to know that it is also related to the peri-implantatory processes that involve the sperm-oocyte interaction, cellular changes observed during fertilization, and the early and late embryo development. Several pathways have been involved at the early stages of human gametogenesis. The spermatozoon has demonstrated an intricate correlation during the fertilization process, as a transfected vector on genetic material, and as interacting with other inner components (RNAm, mitochondrial organelles, etc.). Spermatogenesis is affected by programmed death cell pathways from its packaging process through the elongated cytoplasmic structures during spermiogenesis. Flow cytometry (FC) has been an outstanding tool with the capability to select human gametes to achieve a better reproductive condition. It has been applied as a diagnostic and therapeutic tool allowing a measurable and objective selection and discrimination of spermatozoa from subfertile subjects. Using FC, we are able to know that early distribution of organelles such as mitochondria has an impact in embryo quality before genetic activation on the eight-cell stages occurs. This chapter will let the readers know the current knowledge on sperm fertilization and the relation between the embryo development and the offspring and all the tools now available for an early diagnosis and to identify therapeutic options with FC.
关键词： apoptosis,flow cytometry,spermatozoa,sperm,DNA fragmentation,embryo development,fertilization
更新于2025-09-09 09:28:46
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Synthesized glucocorticoid-induced leucine zipper peptide (GILZ-P) inhibits photoreceptor apoptosis and protects retinal function in light-induced retinal degeneration model
摘要： Background: This study aimed to investigate the neuroprotective function of a synthesized glucocorticoid-induced leucine zipper peptide (GILZ-p) in a light-induced retinal degeneration model. Methods: The GILZ98–134 peptide was synthesized and injected intravitreally into Sprague Dawley rats. Retinal injury was then induced in the rats by exposing their eyes to constant white light (5000 lux) for 24 h. The activation of retinal caspases-9/3 and the release of cytochrome c from the mitochondria to the cytosol were measured at 1, 3, 5 and 7 days after light injury. Photoreceptor apoptosis was evaluated with TUNEL staining at 3 days after injury. Hematoxylin and eosin staining and electroretinography were used to observe the changes in the retinal morphology and function, respectively, 7 and 14 days after light injury. Results: The intravitreally injected synthesized GILZ-p successfully penetrated to the retina and significantly inhibited the activation of retinal caspase-3 and caspase-9 at 1, 3, 5, and 7 days after light injury, and reduced the number of TUNEL-positive photoreceptors at 3 days after light injury. GILZ-p pretreatment also alleviated cytochrome c release and rescued mitochondria-mediated apoptosis after injury. Simultaneously, GILP-p pretreatment also mitigated the light-induced thinning of the outer nuclear layer and the loss of retinal function at 7 and 14 days after light injury, respectively. Conclusion: The synthesized GILZ-p prevented light-induced photoreceptor apoptosis and protected retinal function from degeneration, and is therefore a potential therapeutic option for degenerative retinal diseases.
关键词： photoreceptor,retinal degeneration,synthesized peptide,apoptosis,GILZ
更新于2025-09-09 09:28:46
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Effects of TONS504?photodynamic therapy on mouse mammary tumor cells
摘要： In the present study, TONS504 (C51H58N8O5I2; molecular weight, 1,116.9), a novel cationic hydrophilic photosensitizer, was synthesized from protoporphyrin IX dimethyl ester through a five-step process according to a patented method for use in photodynamic therapy (PDT). The subcellular localization of TONS504 and the cytotoxic effects of TONS504-mediated PDT in the mouse mammary tumor EMT6 cell line were investigated. TONS504 was localized primarily in the lysosomes and partially in the mitochondria. The cytotoxic effects of TONS504-mediated PDT in the mouse mammary tumor EMT6 cell line were investigated using a WST8 assay and an Oxidative Stress kit. The cell viability values following treatment with 10 μg/ml TONS504 at light energies of 0, 1, 5 and 10 J/cm2 were 92.5, 101.8, 27.7 and 1.8%, respectively. The percentages of reactive oxygen species (ROS)(+) cells following the same treatment were 8.6, 8.5, 29.2 and 70.1%, respectively, whereas the percentages of apoptotic cells were 7.1, 5.6, 24.8 and 48.7%, respectively. The percentages of ROS(+) and apoptotic cells in the group subjected to TONS504-mediated PDT increased in a manner dependent on the TONS504 concentration and light energy. Further studies are required to evaluate the in vivo pharmacokinetics, tissue distribution and photodynamic effects of TONS504.
关键词： photodynamic therapy,apoptosis,mammary tumor,reactive oxygen species,TONS504
更新于2025-09-09 09:28:46
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Susceptibility of <i>In Vitro</i> Melanoma Skin Cancer to Photoactivated Hypericin versus Aluminium(III) Phthalocyanine Chloride Tetrasulphonate
摘要： The sensitivity of human melanoma cells to photoactivated Hypericin (Hyp) compared to aluminium(III) phthalocyanine chloride tetrasulphonate (AlPcS4Cl) is reported in this study. Melanoma cells (A375 cell line) were treated with various concentrations of Hyp or AlPcS4Cl alone, for 1, 4, and 24 hrs; varying doses of laser irradiation alone (594 or 682 nm); or optimal concentrations of PSs combined with laser irradiation. Changes in cell morphology, viability, membrane integrity, and proliferation after treatment of cells were determined using inverted microscopy, Trypan blue cell exclusion, Lactate Dehydrogenase (LDH) membrane integrity, and adenosine triphosphate (ATP) cell proliferation assay, respectively. More than 60% of cell survival was observed when cells were treated with 2.5 ??M of Hyp or AlPcS4Cl alone at all incubation times or with 5 J/cm2 of 594 or 682 nm laser alone. Combination of PSs and respective lasers leads to a statistically significant incubation time-dependent decrease in survival of cells. Flow cytometry using the FITC Annexin V/PI apoptosis kit demonstrated that cell death induced after Hyp-PDT is via early and late apoptosis whereas early apoptosis was the main mechanism observed with AlPcS4Cl-PDT. Hyp-PDT compared to AlPcS4Cl-PDT is indicated to be a more effective cancer cell death inducer in melanoma cells.
关键词： AlPcS4Cl,apoptosis,Hypericin,photodynamic therapy,melanoma
更新于2025-09-09 09:28:46
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Cervical cancer cell lines are sensitive to sub-erythemal UV exposure
摘要： High risk human papillomavirus (HPV) infections are the causative agent in virtually every cervical cancer as well as a host of other anogenital and oropharyngeal malignancies. These viruses must activate DNA repair pathways to facilitate their replication, while avoiding the cell cycle arrest and apoptosis that can accompany DNA damage. HPV oncoproteins facilitate each of these goals, but also reduce genome stability. Our data dissect the cytotoxic and cytoprotective characteristics of HPV oncogenes in cervical cancer cells. These data show that while the transformation of keratinocytes by HPV oncogene leaves these cells more sensitive to UV, the onocogenes also protect against UV-induced apoptosis. Cisplatin and UV resistant cervical cancer cell lines were generated and probed for their sensitivity to genotoxic agents. Cervical cancer cells can acquire resistance to one DNA crosslinking agent (UV or cisplatin) without gaining broad tolerance of crosslinked DNA. Further, cisplatin resistance may or may not result in sensitivity to PARP1 inhibition.
关键词： HPV,cisplatin resistance,apoptosis,cervical cancer,PARP1 inhibition,Human Papillomavirus,DNA repair,UV exposure
更新于2025-09-09 09:28:46