Dead Cells Research Articles

Macrophages in necrotic liver lesion repair: opportunities for therapeutical applications.

Healthy livers contain 80% of body resident macrophages known as Kupffer cells. In diseased livers, the number of Kupffer cells usually drops but is compensated by infiltration of monocyte-derived macrophages, some of which can differentiate into Kupffer-like cells. Early studies suggest that Kupffer cells play important roles in both promoting liver injury and liver regeneration. Yet, the distinction between the functionalities of resident and infiltrating macrophages is not always made. By using more specific macrophage markers and targeted cell depletion and single-cell RNA sequencing, recent studies revealed several subsets of monocyte-derived macrophages that play important functions in inducing liver damage and inflammation as well as in liver repair and regeneration. In this review, we discuss the different roles that hepatic macrophages play in promoting necrotic liver lesion resolution and dead cell clearance, as well as the targeting of these cells as potential tools for the development of novel therapies for acute liver failure and acute-on-chronic liver failure.

Development of Biphasic Injectable Hydrogels for Meniscus Scaffold from Photocrosslinked Glycidyl Methacrylate-Modified Poly(Vinyl Alcohol)/Glycidyl Methacrylate-Modified Silk Fibroin.

The development of a hydrogel material with a modified chemical structure of poly(vinyl alcohol) (PVA) and silk fibroin (SF) using glycidyl methacrylate (GMA) (denoted as PVA-g-GMA and SF-g-GMA) is an innovative approach in the field of biomaterials and meniscus tissue engineering in this study. The PVA-g-GMA/SF-g-GMA hydrogel was fabricated using different ratios of PVA-g-GMA to SF-g-GMA: 100/0, 75/25, 50/50, 25/75, and 0/100 (w/w of dry substances), using lithium phenyl (2,4,6-trimethylbenzoyl)phosphinate (LAP) as a free radical photoinitiator, for 10 min at a low ultraviolet (UV) intensity (365 nm, 6 mW/cm2). The mechanical properties, morphology, pore size, and biodegradability of the PVA-g-GMA/SF-g-GMA hydrogel were investigated. Finally, for clinical application, human chondrocyte cell lines (HCPCs) were mixed into PVA-g-GMA/SF-g-GMA solutions and fabricated into hydrogel to study the viability of live and dead cells and gene expression. The results indicate that as the SF-g-GMA content increased, the compressive modulus of the PVA-g-GMA/SF-g-GMA hydrogel dropped from approximately 173 to 11 kPa. The degradation rates of PVA-g-GMA/SF-g-GMA 100/0, 75/25, and 50/50 reached up to 15.61%, 17.23%, and 18.93% in 4 months, respectively. In all PVA-g-GMA/SF-g-GMA conditions on day 7, chondrocyte cell vitality exceeded 80%. The PVA-g-GMA/SF-g-GMA 75:25 and 50:50 hydrogels hold promise as a biomimetic biphasic injectable hydrogel for encapsulated augmentation, offering advantages in terms of rapid photocurability, tunable mechanical properties, favorable biological responses, and controlled degradation.

Unraveling the Anti-Aging Properties of Phycocyanin from the Cyanobacterium Spirulina (Arthrospira platensis).

In recent years, marine natural products have become one of the most important resources of novel lead compounds for critical diseases associated with age. Spirulina, a dietary supplement made from blue-green algae (cyanobacteria: scientific name Arthrospira platensis), is particularly rich in phycocyanin, a phycobiliprotein, which accounts for up to 20% of this cyanobacterium's dry weight and is considered responsible for its anti-cancer, anti-inflammatory and antioxidant activities. Although the anti-aging activity of phycocyanin has been investigated, how exactly this compound works against aging remains elusive. The aim of our research is to use the yeast Saccharomyces cerevisiae as a model organism to investigate the anti-aging properties of phycocyanin from A. platensis. Our results show that phycocyanin has a powerful anti-aging effect, greatly extending the chronological life span of yeast cells in a dose-dependent way, as the effect was also pronounced when cells were grown in SD medium under calorie restriction conditions (0.2% glucose). Both ROS and accumulation of dead cells were followed by staining chronologically aged cells with dihydrorhodamine 123 (DHR123) and propidium iodide (PI). Interestingly, we found that most of the aged phycocyanin-treated cells, which were unable to form colonies, were actually ROS+/PI-. Finally, we show that the moment in which phycocyanin is added to the culture does not substantially influence its effectiveness in counteracting chronological aging.

Microalgal cell division tracking using CFSE

Microalgal cell division tracking could unlock new research means. It could help decipher cell response to an intentional stressor, aiming at increasing an added value molecule accumulation, or an accidental stressor, in the case of environmental pollution. It could also be used to monitor asynchrony in cultures exposed to photoperiod or determine the fate of dead cells. To date, because of the lack of guidelines specific to microalgae, microalgal ecotoxicology and biotechnology communities have not yet implemented such protocol in routine. Therefore, a systematic optimization methodology has been deployed to adapt the CarboxyFluorescein DiAcetate Succinimidyl Ester (CF-DA-SE, or, in short, CFSE) lymphocytes proliferation tracking technique to the microalga Chlorella vulgaris. The toxic effect of the CFSE solvent (DMSO) was delineated (stock solution at 10 mM). Then, incubation conditions (time, probe/cell ratio, illumination) were optimized (30 min, 4.50 nmol/MCell, in the dark). Finally, CFSE washing and cell recovery were robustified (low-acceleration - 100 g - centrifugation). Using a semi-synchronous culture as a test case, the method was successfully applied to count cell divisions. Up to four generations could be discriminated. The generation-to-generation signal ratio was exactly 1/4, corresponding to the natural division of Chlorella vulgaris. Furthermore, an advanced yet easy-to-implement signal processing technique was introduced to ease generation discrimination.

Fluorogenic Chemical Probes for Wash-free Imaging of Cell Membrane Damage in Ferroptosis, Necrosis, and Axon Injury.

Selectively labeling cells with damaged membranes is needed not only for identifying dead cells in culture, but also for imaging membrane barrier dysfunction in pathologies in vivo. Most membrane permeability stains are permanently colored or fluorescent dyes that need washing to remove their non-uptaken extracellular background and reach good image contrast. Others are DNA-binding environment-dependent fluorophores, which lack design modularity, have potential toxicity, and can only detect permeabilization of cell volumes containing a nucleus (i.e., cannot delineate damaged volumes in vivo nor image non-nucleated cell types or compartments). Here, we develop modular fluorogenic probes that reveal the whole cytosolic volume of damaged cells, with near-zero background fluorescence so that no washing is needed. We identify a specific disulfonated fluorogenic probe type that only enters cells with damaged membranes, then is enzymatically activated and marks them. The esterase probe MDG1 is a reliable tool to reveal live cells that have been permeabilized by biological, biochemical, or physical membrane damage, and it can be used in multicolor microscopy. We confirm the modularity of this approach by also adapting it for improved hydrolytic stability, as the redox probe MDG2. We conclude by showing the unique performance of MDG probes in revealing axonal membrane damage (which DNA fluorogens cannot achieve) and in discriminating damage on a cell-by-cell basis in embryos in vivo. The MDG design thus provides powerful modular tools for wash-free in vivo imaging of membrane damage, and indicates how designs may be adapted for selective delivery of drug cargoes to these damaged cells: offering an outlook from selective diagnosis toward therapy of membrane-compromised cells in disease.

In Vitro Investigation of the Antimicrobial Potential and Cell Viability of Different Dosages of Ozone Therapy

ABSTRACT The aim of the study was to evaluate and compare, in vitro, the cytotoxicity of different applied ozone dosages in keratinocyte (NOK) and fibroblast (L929) cell lines, as well as the antimicrobial potential for Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli). NOK and L929 cells and strains of gram-positive bacteria S. aureus and gram-negative bacteria E. coli were divided into seven experimental groups: ozone at doses from 0, 0.005, 0.01, 0.02, 0.03, 0.04 and 0.05 mg/L. The effect of ozone on cell viability/cytotoxicity was determined using the alamarBlue® assay, the Live/Dead® assay and the morphometric analysis of the percentage of live and dead cells. The results show that applied ozone dosages of 0.005, 0.01 and 0.02 mg/L showed the highest viability values of NOK and L929 cells, as well as a higher density and percentage of live cells in the 72-hour experimental. However, no difference was observed in the percentage of dead cells between the experimental groups. Furthermore, ozone promoted a reduction in CFUs at all tested dosages. With the results obtained in the present study, it is possible to confirm the restorative potential of ozone therapy in a dose-dependent response, as well as its antimicrobial potential.

Evaluation of the antimicrobial and antioxidant properties of synthetic phenolipids

Functionalization of phenolic compounds by lipophilization allowing the synthesis of new compounds with improved biological properties. This study aims to investigate the antimicrobial effects of eight synthesized phenolipids, derived from bioactive coumaric and ferulic acid esters, against Bacillus subtilis, Escherichia coli, and Candida albicans. Antioxidant activity, as well as antimicrobial capacity were evaluated through DPPH assays and classical disc agar diffusion, respectively, concomitantly with the evaluation of cellular viability using flow cytometry assay. Antioxidant activities results showed that ferulic acid esters exhibit higher antiradical activity than coumaric acid esters. Results of agar diffusion assay revealed that, 1-methylpropyl coumarate and 1-methylpropyl ferulate (1-MPF) have an effective antimicrobial activity with Minimum Inhibitory Concentrations (MICs) values between 0.9 and 3.5 mM against all tested microorganisms. Also, flow cytometry showed that 1-MPF and decyl coumarate exhibited significant bactericidal activity (p < 0.05), with almost 63 % of dead Bacillus subtilis cells. Compared to the commonly used agar method, flow cytometry proved to be a fast and straightforward analytical tool to assess antimicrobial activity while providing information on inhibition type. These results suggest the applicability of phenolipids as potent lipophilic bioactive molecules to protect food products and cosmetic formulations from oxidative stress and microbial spoilage.

FORMATION OF NANOCONES AND GENERATION OF NEGATIVE POTENTIAL ON STAINLESS STEEL SURFACES BY ELECTROCHEMICAL ETCHING SYNERGISTICALLY REDUCE PSEUDOMONAS AERUGINOSA'S BIOFILM

Hospital-acquired infections cause severe patient problems because of the augmented appearance of antibiotic-resistant bacteria, including Pseudomonas aeruginosa. Material surfaces modified with several biophysical parameters can decrease bacterial biofilm formation, which could be an advantageous alternative to treatment with antibiotics. Since stainless steel is an extensively used material for manufacturing medical implants and in healthcare settings, in this study, we used stainless steel (SS 316L and SS 304) to examine the result of the material surface topographies on bacterial biofilm establishment. This work used the electrochemical etching method to modify the stainless steel surface topography as an anode. The electrochemical etching method influenced the nanocones’ formation on stainless steel surfaces of both SS316L (Disk-6: 2682 peaks/[Formula: see text]m[Formula: see text] and SS304 (Disk-12: 1654 peaks/[Formula: see text]m[Formula: see text] estimated by atomic force microscopy and 3D Profilometer reduced the biofilm by 78% and 85%, respectively. Additionally, the higher negative potential on an average of 600[Formula: see text]mV measured by Kelvin probe atomic force microscopy reduced the biofilm formation on both SS316L and SS304 surface synergistically compared to the non-electrochemically etched surface. Biofilm growth and nanopotrusions on the stainless-steel surface examined by atomic force microscopy and scanning electron microscopy demonstrated significantly dead bacterial cells (20%) on the electrochemically etched surface than on the non-electrochemically etched surface after 2[Formula: see text]h contact time. Our observations exhibit that the nanotextured surface topographies and surface negative potential effectively inhibit bacterial adhesion and biofilm formation.

Overnutrition and Lipotoxicity: Impaired Efferocytosis and Chronic Inflammation as Precursors to Multifaceted Disease Pathogenesis.

Overnutrition, driven by the consumption of high-fat, high-sugar diets, has reached epidemic proportions and poses a significant global health challenge. Prolonged overnutrition leads to the deposition of excessive lipids in adipose and non-adipose tissues, a condition known as lipotoxicity. The intricate interplay between overnutrition-induced lipotoxicity and the immune system plays a pivotal role in the pathogenesis of various diseases. This review aims to elucidate the consequences of impaired efferocytosis, caused by lipotoxicity-poisoned macrophages, leading to chronic inflammation and the subsequent development of severe infectious diseases, autoimmunity, and cancer, as well as chronic pulmonary and cardiovascular diseases. Chronic overnutrition promotes adipose tissue expansion which induces cellular stress and inflammatory responses, contributing to insulin resistance, dyslipidemia, and metabolic syndrome. Moreover, sustained exposure to lipotoxicity impairs the efferocytic capacity of macrophages, compromising their ability to efficiently engulf and remove dead cells. The unresolved chronic inflammation perpetuates a pro-inflammatory microenvironment, exacerbating tissue damage and promoting the development of various diseases. The interaction between overnutrition, lipotoxicity, and impaired efferocytosis highlights a critical pathway through which chronic inflammation emerges, facilitating the development of severe infectious diseases, autoimmunity, cancer, and chronic pulmonary and cardiovascular diseases. Understanding these intricate connections sheds light on potential therapeutic avenues to mitigate the detrimental effects of overnutrition and lipotoxicity on immune function and tissue homeostasis, thereby paving the way for novel interventions aimed at reducing the burden of these multifaceted diseases on global health.

Antibacterial Activity Of Clay Mask From Houttuynia Cordata Leaves Extract Against Staphylococcus Aureus

Facial skin that often looks dull and not glowing can be caused by the accumulation of dead skin cells. The solution to this problem is to exfoliate using a scrub. The content of secondary metabolites in Amis-amisan leaves (Houttuynia cordata Thunb) has antibacterial activity. This study aimed to determine the antibacterial activity of the face scrub ethanol extract of amis-amisan leaves against Staphylococcus aureus. The extract was obtained using the remaceration method using 96% ethanol. Concentrations of face scrub are 10%, 20%, and 30%. Antibacterial activity using the pour plate method. The results antibacterial activity of face scrub with concentrations of 10%, 20%, and 30% are 1.127 cm, 1.282 cm and 1.460 cm, respectively. The best antibacterial activity is a 30% concentration face scrub. The physical characteristics of the organoleptic, homogeneity, pH, spreadability, adhesion, and irritation tests of the face scrub meet the specified requirements.

Mitochondrial Calcium Uniporter (MCU) is Involved in an Ischemic Postconditioning Effect Against Ischemic Reperfusion Brain Injury in Mice

The phenomenon of ischemic postconditioning (PostC) is known to be neuroprotective against ischemic reperfusion (I/R) injury. One of the key processes in PostC is the opening of the mitochondrial ATP-dependent potassium (mito-KATP) channel and depolarization of the mitochondrial membrane, triggering the release of calcium ions from mitochondria through low-conductance opening of the mitochondrial permeability transition pore. Mitochondrial calcium uniporter (MCU) is known as a highly sensitive transporter for the uptake of Ca2+ present on the inner mitochondrial membrane. The MCU has attracted attention as a new target for treatment in diseases, such as neurodegenerative diseases, cancer, and ischemic stroke. We considered that the MCU may be involved in PostC and trigger its mechanisms. This research used the whole-cell patch-clamp technique on hippocampal CA1 pyramidal cells from C57BL mice and measured changes in spontaneous excitatory post-synaptic currents (sEPSCs), intracellular Ca2+ concentration, mitochondrial membrane potential, and N-methyl-d-aspartate receptor (NMDAR) currents under inhibition of MCU by ruthenium red 265 (Ru265) in PostC. Inhibition of MCU increased the occurrence of sEPSCs (p = 0.014), NMDAR currents (p < 0.001), intracellular Ca2+ concentration (p < 0.001), and dead cells (p < 0.001) significantly after reperfusion, reflecting removal of the neuroprotective effects in PostC. Moreover, mitochondrial depolarization in PostC with Ru265 was weakened, compared to PostC (p = 0.004). These results suggest that MCU affects mitochondrial depolarization in PostC to suppress NMDAR over-activation and prevent elevation of intracellular Ca2+ concentrations against I/R injury.Graphical

Vesicle formation-related protein CaSec16 and its ankyrin protein partner CaANK2B jointly enhance salt tolerance in pepper

Vesicle transport plays important roles in plant tolerance against abiotic stresses. However, the contribution of a vesicle formation related protein CaSec16 (COPII coat assembly protein Sec16-like) in pepper tolerance to salt stress remains unclear. In this study, we report that the expression of CaSec16 was upregulated by salt stress. Compared to the control, the salt tolerance of pepper with CaSec16-silenced was compromised, which was shown by the corresponding phenotypes and physiological indexes, such as the death of growing point, the aggravated leaf wilting, the higher increment of relative electric leakage (REL), the lower content of total chlorophyll, the higher accumulation of dead cells, H2O2, malonaldehyde (MDA), and proline (Pro), and the inhibited induction of marker genes for salt-tolerance and vesicle transport. In contrast, the salt tolerance of pepper was enhanced by the transient overexpression of CaSec16. In addition, heterogeneously induced CaSec16 protein did not enhance the salt tolerance of Escherichia coli, an organism lacking the vesicle transport system. By yeast two-hybrid method, an ankyrin protein, CaANK2B, was identified as the interacting protein of CaSec16. The expression of CaANK2B showed a downward trend during the process of salt stress. Compared with the control, pepper plants with transient-overexpression of CaANK2B displayed increased salt tolerance, whereas those with CaANK2B-silenced exhibited reduced salt tolerance. Taken together, both the vesicle formation related protein CaSec16 and its interaction partner CaANK2B can improve the pepper tolerance to salt stress.

Development of alginate-collagen interpenetrating network for osteoarthritic cartilage by in situ softening

This study presents an alginate-collagen interpenetrating network (IPN) matrix of incorporating collagen fibrils into an alginate hydrogel by physical mixing and controlled gelation. The resulting matrix closely mimics the physiological and pathological stiffness range of the chondrocyte pericellular matrix (PCM). Chondrocytes were cultured within three-dimensional (3D) alginate-collagen IPN matrices with varying stiffness, namely Firm, Medium, and Soft. Alginate lyase was introduced to study the effects of the changes in stiffness of the Firm on chondrocyte response by in situ softening. The developed alginate-collagen IPN matrix displayed good cell-biocompatibility. Compared with stiffer tissue culture plastic (TCP), chondrocytes grown within Firm displayed a stabilized differentiated phenotype characterized by higher expression levels of aggrecan, collagen II, and SOX-9. Moreover, the developed alginate-collagen IPN matrix exhibited a gradually increased percentage of propidium iodide (PI)-positive dead cells with decreasing stiffness. Softer matrices directed cells towards higher proliferation rates and spherical morphologies while stimulating chondrocyte cluster formation. Furthermore, reducing Firm stiffness by in situ softening decreased aggrecan expression, contributing to matrix degradation similar to that seen in osteoarthritis (OA). Hence, the 3D alginate-collagen IPN constructs hold significant potential for in vitro replicating PCM stiffness changes observed in OA cartilage.

Viability and Proliferation Assessment of Gingival Fibroblasts Cultured on Silver Nanoparticle-Doped Ti-6Al-4V Surfaces.

To investigate the biocompatibility of silver nanoparticle (AgNP)-doped Ti-6Al-4V surfaces by evaluating the viability and proliferation rate of human gingival fibroblasts (HGFs)-as the dominant cells of peri-implant soft tissues-seeded on the modified surfaces. AgNPs (sizes 8 nm and 30 nm) were incorporated onto Ti-6Al-4V specimen surfaces via electrochemical deposition, using colloid silver dispersions with increasing AgNP concentrations of 100 ppm, 200 ppm, and 300 ppm. One control and six experimental groups were included in the study: (1) control (Ti-6Al-4V), (2) 8 nm/100 ppm, (3) 8 nm/200 ppm, (4) 8 nm/300 ppm, (5) 30 nm/100 ppm, (6) 30 nm/200 ppm, and (7) 30 nm/300 ppm. HGF cell primary cultures were isolated from periodontally healthy donor patients and cultured in direct contact with the group specimens for 24 and 72 hours. The cytotoxicity of AgNP-doped Ti-6Al-4V specimens toward HGF was assessed by the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) and BrdU (5-bromo-2'-deoxyuridine) assay tests. Calcein AM and ethidium homodimer (EthD-1) fluorescent stains were used to determine the live and dead cells. The morphology and attachment properties of the HGFs were determined via scanning electron microscopy (SEM). Energy dispersive x-ray (EDX) analysis confirmed the presence of AgNPs on the specimens. The MTT test revealed that AgNPs of both sizes and all concentrations presented a decreased cellular metabolic activity compared to the control discs. All concentrations of both sizes of AgNPs affected the cell proliferation rate compared to the control group, as revealed by the BrdU assay. Overall, cytotoxicity of the modified Ti-6Al-4V surfaces depended on cell exposure time. Observation via confocal microscopy confirmed the results of the MTT and BrdU assay tests. Specifically, most cells remained alive throughout the 72-hour culture period. SEM images revealed that adjacent cells form bonds with each other, creating confluent layers of conjugated cells. The findings of the present study indicate that Ti-6Al-4V surfaces modified with 8 nm and 30 nm AgNPs at concentrations of 100 ppm, 200 ppm, and 300 ppm do not produce any serious cytotoxicity toward HGFs. The initial arrest of the HGF proliferation rate recovered at 72 hours. These results on the antibacterial activity against common periodontal pathogens, in combination with the results found in a previous study by the same research group, suggest that AgNP-doped Ti-6Al-4V surfaces are potential candidates for use in implant abutments for preventing peri-implant diseases.

Advances on extracellular vesicles derived from dead cells

Advances on extracellular vesicles derived from dead cells

Electrochemical Assay of Mammalian Cell Viability

We present the first use of amperometric detection to assess the viability of mammalian cells in continuous mode, directly in the cell culture medium. Vero or HeLa cells were injected into electrochemical sensors equipped with a 3-electrode system and containing DCIP 50 µM used as the redox mediator. DCIP was reduced by the viable cells and the reduced form was detected amperometrically at 300 mV vs silver pseudo-reference. The continuous regeneration of the oxidized form of the mediator ensured a stable redox state of the cell environment, allowing the cells to survive during the measurement time. The electrochemical response was related to cell metabolism (no response with dead cells or lysed cells) and depended on both mediator concentration and cell density. The protocol was applied to both cells in suspension and adhered cells. It was also adapted to detect trans-plasma membrane electron transfer (tPMET) by replacing DCIP by ferricyanide 500 µM and using linear scan voltammetry (2 mV/s). The pioneering results described here pave the way to the development of routine electrochemical assays for cell viability and for designing a cell-based analytical platform.

Effectiveness of Pheophorbide-a mediated Photodynamic Therapy against Doxorubicin Resistant Breast Cancer

SignificanceEmploying mechanistically distinct therapies like Photodynamic Therapy (PDT) can be a promising strategy to overcome the multidrug- resistance in breast cancer resulting in the killing of chemotherapeutic drug-resistant cancer cells. ApproachIn this study, Pheophorbide a (Pheo-a), is being used as a potential photosensitizer to induce cell death in Doxorubicin (DOX) resistant breast cancer MCF-7 cells. Efficacy of Pheo-a mediated PDT against MCF-7/DOX cancer cells was evaluated by MTT proliferation assay and analyzing cellular morphological changes, live dead cell death assay. ResultsAll experiments were performed thrice in triplicate, and the results obtained were analysed for statistical significance. Results showed that the Pheo-a can induce photodynamic cell death in resistant cancer cells treated with a 2.5 μM concentration of Pheo-a followed by irradiation with a 660 nm red laser at a light dose of 10 J/cm2. ConclusionsThus, the results from this study suggest the utilization of chlorophyll-based PSs as a rational therapeutic intervention for the eradication of chemo drug-resistance in cancers.

Short Communication: Mammary epithelial cell exfoliation increases as milk yield declines, lactation progresses, and parity increases.

Milk yield (MY) naturally declines after peak lactation. One approach to improving production is to increase lactation persistency, or the length of time a cow maintains lactation. To do so, cows must maintain functional secretory mammary epithelial cells (MEC), which synthesize milk. Some viable MEC are exfoliated from the udder into milk. The objective of this experiment was to identify the magnitude and direction of association between exfoliated MEC and MY, days in milk (DIM), and parity. Our hypothesis was that MY and the quantity of exfoliated MEC were negatively correlated. Therefore, milk from low yielding, late lactation, and primiparous cows would contain more MEC. Thirty-six multiparous and 12 primiparous Holstein cows were sampled once during lactation. A single milk sample was obtained from each cow during a morning milking. Flow cytometry was used to identify milk cell types. Cells were stained with an antibody specific for cluster of differenciation-45 (CD45) to label hematopoietic cells and exclude them from analysis. An antibody against butyrophilin 1A1 (BTN), a protein involved in milk fat secretion, labeled MEC. Nucleated cells were stained with Hoechst dye, and propidium iodide was used to identify dead cells. For statistical analysis, we analyzed the effect of parity, MY, and DIM, and DIM2 on the number and concentration of both total and live exfoliated MEC. Primiparous cows exfoliated fewer MEC and had a lower concentration of MEC in milk than multiparous cows regardless of lactation stage. This could reflect continued mammary growth in first lactation. Cows shed an average of 9.80x107 ± 6.907 MEC with a concentration of 4,582 ± 3,574 cells/mL in one milking. Milk yield was inversely related to concentration of both total and live MEC in milk. Similarly, cows in late lactation, which is associated with MY decline, exfoliated a greater number of MEC into milk. Primiparous cows had a lower milk yield and shed fewer total and live MEC in this study. The relationship of MY and DIM with MEC losses in milk may also contribute to declining MEC number in the mammary gland found in post-peak lactation. Understanding MEC exfoliation regulation could help develop husbandry practices that improve secretory cell retention in the mammary gland and increase lactation persistency.

Zinc oxide nanoparticles preserve the quality and fertility potential of rooster sperm during the cryopreservation process.

Sperm cryopreservation is one of the main methods for preserving rooster sperm for artificial insemination (AI) in commercial flocks. Yet, rooster sperm is extremely susceptible to reactive oxygen species (ROS) produced during the freezing process. Oxidative stress could be prevented by using nanoparticles containing antioxidants. The present study was conducted to investigate the effect of zinc oxide nanoparticles (ZnONP) in rooster semen freezing extender on quality parameters and fertility potential. For this aim, semen samples were collected and diluted in Lake extenders as follows: control: Lake without ZnONP, ZnO100: Lake with 100-μg zinc oxide (ZnO), ZnONP50: Lake with 50-μg ZnONP, ZnONP100: Lake with 100-μg ZnONP and ZnONP200: Lake with 200-μg ZnONP. After freezing and thawing, sperm motility, viability, membrane integrity, morphology, mitochondrial activity, acrosome integrity, DNA fragmentation, lipid peroxidation and ROS, as well as fertility and hatchability were assessed. According to the current results, higher rates of motility, membrane integrity, mitochondrial activity, acrosome integrity and live cells were detected in the ZnO100, ZnONP50 and ZnONP100 groups compared to other groups (p ≤ .05). Yet, the percentage of dead cells, DNA fragmentation, lipid peroxidation and ROS levels were lower in the mentioned groups (p ≤ .05). Furthermore, a higher percentage of fertility was observed in the ZnO100 and ZnONP100 groups than in the control group (p ≤ .05). In conclusion, the use of 100-μg ZnO and 50- to 100-μg ZnONP represents a valuable and safe additive material that could be used to improve the quality and fertility potential of rooster sperm under cryopreservation conditions.

Effectiveness of Pheophorbide-a mediated Photodynamic Therapy against Doxorubicin Resistant Breast Cancer

SignificanceEmploying mechanistically distinct therapies like Photodynamic Therapy (PDT) can be a promising strategy to overcome the multidrug- resistance in breast cancer resulting in the killing of chemotherapeutic drug-resistant cancer cells. ApproachIn this study, Pheophorbide a (Pheo-a), is being used as a potential photosensitizer to induce cell death in Doxorubicin (DOX) resistant breast cancer MCF-7 cells. Efficacy of Pheo-a mediated PDT against MCF-7/DOX cancer cells was evaluated by MTT proliferation assay and analyzing cellular morphological changes, live dead cell death assay. ResultsAll experiments were performed thrice in triplicate, and the results obtained were analysed for statistical significance. Results showed that the Pheo-a can induce photodynamic cell death in resistant cancer cells treated with a 2.5 μM concentration of Pheo-a followed by irradiation with a 660 nm red laser at a light dose of 10 J/cm2. ConclusionsThus, the results from this study suggest the utilization of chlorophyll-based PSs as a rational therapeutic intervention for the eradication of chemo drug-resistance in cancers.