Human Cells Research Articles

Complexation by γ-cyclodextrin as a way of improving anticancer potential of sumanene

Abstract Biological applications of sumanene buckybowl molecule have been widely discussed over the years yet remain still unexplored experimentally. On the other hand, creating cyclodextrin-containing supramolecular assemblies was demonstrated to be a powerful tool in terms of designing effective systems for medicinal chemistry purposes. Here, we show that sumanene molecule exclusively forms 1:1 host-guest complexes with γ-cyclodextrin (γCD) or (2-hydroxypropyl)-γ-cyclodextrin (HP-γCD), as revealed by extensive spectroscopic studies supported with density functional theory (DFT) computations. Based on our preliminary biological studies, we discovered that the formation of such complexes resulted in the improvement of anticancer properties of sumanene, expressed by high cell viabilities in vitro of healthy human mammary fibroblasts (HMF) together with low viabilities of human breast adenocarcinoma cells (MDA-MB-231). Improved pharmacokinetic (ADME-Tox) properties for sumanene@γCD and sumanene@HP-γCD complexes in comparison to native sumanene were also supported by in sillico modeling studies. This work provides the method how to focus the cytotoxic action of sumanene toward cancer cells using supramolecular assembly strategy, paving the way to the further exploration of biological properties of sumanene-containing supramolecular systems with bioactive features and applications of this buckybowl in general.

Application of RET Approach for Ratiometric Response Enhancement of ICT Fluorescent Hg2+ Probe based on Crown‐containing Styrylpyridinium Dye

AbstractStyrylpyridinium dye bearing azadithia‐15‐crown‐5 ether receptor group SP and 4‐alkoxy‐1,8‐naphthalimide fluorophore were linked using copper‐catalyzed azide‐alkyne cycloaddition click reaction to afford dyad compound NI‐SP. Chemosensor NI‐SP exhibited selective ratiometric fluorescent response to the presence of Hg2+ in aqueous solution due to the interplay between resonance energy transfer (RET) and intramolecular charge transfer (ICT) processes occurred upon excitation. The observed switching of the ratio of emission intensities in the blue and red channels R was higher than in the case of monochromophoric styrylpyridine derivative SP showing ratiometric response based on ICT mechanism only. Biological studies revealed that NI‐SP penetrates into human lung adenocarcinoma A549 cells and accumulates in cytoplasm and lysosomes. When cells were pre‐incubated with mercury (II) perchlorate, the ratio R was increased 2.6 times, which enables detection of intracellular Hg2+ ions and their quantitative analysis in the 0.7–6.0 μM concentration range.

Epigenetic dynamics of partially methylated domains in human placenta and trophoblast stem cells.

The placenta is essential for nutrient exchange and hormone production between the mother and the developing fetus and serves as an invaluable model for epigenetic research. Most epigenetic studies of the human placenta have used whole placentas from term pregnancies and have identified the presence of partially methylated domains (PMDs). However, the origin of these domains, which are typically absent in most somatic cells, remains unclear in the placental context. Using whole-genome bisulfite sequencing and analysis of histone H3 modifications, we generated epigenetic profiles of human cytotrophoblasts during the first trimester and at term, as well as human trophoblast stem cells. Our study focused specifically on PMDs. We found that genomic regions likely to form PMDs are resistant to global DNA demethylation during trophectoderm reprogramming, and that PMDs arise through a slow methylation process within condensed chromatin near the nuclear lamina. In addition, we found significant differences in histone H3 modifications between PMDs in cytotrophoblasts and trophoblast stem cells. Our findings suggest that spatiotemporal genomic features shape megabase-scale DNA methylation patterns, including PMDs, in the human placenta and highlight distinct differences in PMDs between human cytotrophoblasts and trophoblast stem cells. These findings advance our understanding of placental biology and provide a basis for further research into human development and related diseases.

Coordinated inheritance of extrachromosomal DNAs in cancer cells.

The chromosomal theory of inheritance dictates that genes on the same chromosome segregate together while genes on different chromosomes assort independently1. Extrachromosomal DNAs (ecDNAs) are common in cancer and drive oncogene amplification, dysregulated gene expression and intratumoural heterogeneity through random segregation during cell division2,3. Distinct ecDNA sequences, termed ecDNA species, can co-exist to facilitate intermolecular cooperation in cancer cells4. How multiple ecDNA species within a tumour cell are assorted and maintained across somatic cell generations is unclear. Here we show that cooperative ecDNA species are coordinately inherited through mitotic co-segregation. Imaging and single-cell analyses show that multiple ecDNAs encoding distinct oncogenes co-occur and are correlated in copy number in human cancer cells. ecDNA species are coordinately segregated asymmetrically during mitosis, resulting in daughter cells with simultaneous copy-number gains in multiple ecDNA species before any selection. Intermolecular proximity and active transcription at the start of mitosis facilitate the coordinated segregation of ecDNA species, and transcription inhibition reduces co-segregation. Computational modelling reveals the quantitative principles of ecDNA co-segregation and co-selection, predicting their observed distributions in cancer cells. Coordinated inheritance of ecDNAs enables co-amplification of specialized ecDNAs containing only enhancer elements and guides therapeutic strategies to jointly deplete cooperating ecDNA oncogenes. Coordinated inheritance of ecDNAs confers stability to oncogene cooperation and novel gene regulatory circuits, allowing winning combinations of epigenetic states to be transmitted across cell generations.

A B7-H3–Targeted CD28 Bispecific Antibody Enhances the Activity of Anti–PD-1 and CD3 T-cell Engager Immunotherapies

Abstract T-cell activation is a multistep process requiring T-cell receptor engagement by peptide–MHC complexes (Signal 1) coupled with CD28-mediated costimulation (Signal 2). Tumors typically lack expression of CD28 ligands, so tumor-specific Signal 1 (e.g., neoepitope presentation) without costimulation may be ineffective or even induce T-cell anergy. We designed the bispecific antibody XmAb808 to co-engage the tumor-associated antigen B7-H3 with CD28 to promote T-cell costimulation within the tumor microenvironment. XmAb808 costimulation was measured by its ability to activate and expand T cells and enhance T cell–mediated cancer cell killing in cocultures of human peripheral blood mononuclear cells and cancer cells and in mice engrafted with human peripheral blood mononuclear cells and tumor xenografts. XmAb808 avidly bound cancer cells and stimulated IL2 and IFNγ secretion from T cells cocultured with cancer cells engineered to deliver Signal 1 to T cells via a surface-expressed anti-CD3 antibody. XmAb808 enhanced expression of the antiapoptotic factor Bcl-xL and CD25, promoting survival and IL2-dependent expansion of T cells coupled with increased T cell–mediated cytotoxicity in vitro. XmAb808 combined with an EpCAM×CD3 bispecific antibody to enhance target cell killing through IL2-dependent expansion of CD25+ T cells. This combination also suppressed pancreatic tumor xenograft growth in mice. Furthermore, XmAb808 combined with an anti–programmed cell death protein 1 antibody to suppress breast tumor xenograft growth in mice. XmAb808 as monotherapy and in combination with an anti–programmed cell death protein 1 antibody is currently in clinical development in patients with advanced solid tumors. Our results suggest that XmAb808 may also combine with tumor antigen–targeted anti-CD3 (Signal 1) T-cell engagers.

CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells.

We investigated the regulation of histone deacetylases (HDACs) by miR-2861 in the osteoblastic differentiation of human mesenchymal stem cells (MSCs) and miR-2861 binding site by CRISPR activation (CRISPRa). Transfection of miR-2861 into human MSCs was performed and the effect on osteoblast differentiation was analyzed. Using catalytically inactive Cas12a, the CRISPRa system induced targeted overexpression of endogenous miRNA and repressed the luciferase activities of reporters that contained functional miRNA target sites. The delivery of miR-2861 into MSCs enhanced osteoblast differentiation by decreased expressions of the HDAC1, 4 and 5 genes. The mechanism of HDAC5 repression by miR-2861 in humans has not been fully elucidated. To this end, the HDAC5 mRNA sequence was analyzed and a putative primate-specific miR-2861 binding site was identified in the 3' untranslated region (3'-UTR). CRISPRa was applied to validate the putative binding site and an increase in endogenous miR-2861 was found to repress the expression of a reporter that contained the novel miR-2861 binding site. The delivery of miR-2861 to human MSCs enhanced osteoblast differentiation. In the 3'-UTR, the HDAC5 repression was mediated by the miR-2861 binding site, and miR-2861 promoted osteoblast differentiation via the inhibition of HDAC5 through a primate-specific miRNA binding site. Therefore, miRNAmiR-2861 with the CRISPRa methods might be a good biomaterial for osteogenesis augmentation.

Novel Gene Biomarkers Specific to Human Mesenchymal Stem Cells Isolated from Bone Marrow

In this paper, we present a comparative analysis of the transcriptomic profile of three different human cell types: hematopoietic stem cells (HSCs), bone marrow-derived mesenchymal stem cells (MSCs) and fibroblasts (FIBs). The work aims to identify unique genes that are differentially expressed as specific markers of bone marrow-derived MSCs, and to achieve this undertakes a detailed analysis of three independent datasets that include quantification of the global gene expression profiles of three primary cell types: HSCs, MSCs and FIBs. A robust bioinformatics method, called GlobalTest, is used to assess the specific association between one or more genes expressed in a sample and the outcome variable, that is, the ‘cell type’ provided as a single univariate response. This outcome variable is predicted for each sample tested, based on the expression profile of the specific genes that are used as input to the test. The precision of the tests is calculated along with the statistical sensitivity and specificity for each gene in each dataset, yielding four genes that mark MSCs with high accuracy. Among these, the best performer is the protein-coding gene Transgelin (TAGLN, Gene ID: 6876) (with a Positive Predictive Value > 0.96 and FDR < 0.001), which identifies MSCs better than any of the currently used standard markers: ENG (CD105), THY1 (CD90) or NT5E (CD73). The results are validated by RT-qPCR, providing novel gene biomarkers specific for human MSCs.

Novel Schiff bases of quinolin-4(1h)-one: Synthesis, antiproliferative evaluation, apoptosis, cell cycle, autophagy and molecular docking studies in human colon cancer cells

A green and efficient ultrasound-mediated protocol was developed for the synthesis of a series of novel Schiff bases derived from quinoline-4-one. The synthetic strategy involved the condensation of 4-oxo-1,4-dihydro-quinoline-3-carboxaldehyde with diverse anilines and heteroaryl amines. The synthesized compounds were characterized using spectroscopic techniques. A comprehensive in vitro cytotoxicity evaluation against HCT116 and HT-29 human colon cancer cell lines revealed that several compounds exhibited potent anticancer activity. Notably, compounds 3c and 3e demonstrated superior cytotoxicity compared to the clinical standard doxorubicin. Mechanistic studies indicated that these lead compounds induced apoptosis, necrosis, and cell cycle arrest at G1 and G2 phases. Furthermore, autophagy induction was observed. In silico ADMET predictions support the potential of compounds 3c and 3e as promising anticancer drug candidates. The molecular docking studies revealed that the Schiff bases 3c and 3e displayed good interaction with VEGFR-2 receptor. These findings underscore the quinoline-4-one scaffold as a valuable template for the development of novel antitumor agents.

Preparation of DOX-TPP/HA-ss-OA Nanoparticles, Investigation of Drug Release Behavior In Vitro, and Evaluation of Anti-proliferative Activity In Vitro.

This study aimed to develop and characterize DOX-TPP/HA-ss-OA nanoparticles, utilizing the mitochondria-targeting prodrug doxorubicin-triphenylphosphine (DOXTPP) and a reduction-sensitive amphiphilic polymer, hyaluronic acid-disulfide-oleic acid (HAss- OA). The research focused on evaluating the drug release behavior of these nanoparticles under varying glutathione (GSH) concentrations and their anti-tumor activity in vitro. DOX-TPP/HA-ss-OA nanoparticles were prepared using probe ultrasound technology. The study examined the impact of different organic solvents on drug loading capacity and encapsulation efficiency to determine the optimal conditions. A single-factor experimental design was used to optimize the formulation process. Key parameters, including particle size and zeta potential, were measured to assess nanoparticle stability and performance. The dynamic dialysis method was employed to evaluate the reduction-sensitive drug release characteristics in media with different GSH concentrations. The MTT assay was used to analyze the growth-inhibitory effects of the nanoparticles on human breast cancer cells (MCF-7) and drug-resistant cells (MCF-7/ADR). The optimized preparation process for DOX-TPP/HA-ss-OA nanoparticles included a drug dosage of 2.0 mg, an oil-to-water volume ratio of 1:5, ultrasonic power of 500 W, and ultrasonic time of 15 minutes. The nanoparticles had an average particle size of 203.72 ± 2.30 nm and a zeta potential of 25.82 ± 0.58 mV, indicating favorable stability and effective drug delivery properties. The nanoparticles exhibited a slow, sustained release of DOX-TPP in pH 7.4 phosphate buffer solution (PBS) and accelerated release in high GSH concentrations, demonstrating reduction-responsive drug release. In vitro studies showed that DOX-TPP/HA-ss-OA nanoparticles significantly inhibited the proliferation of MCF-7 and MCF-7/ADR cells in a dosedependent manner, with enhanced efficacy compared to free DOX and other formulations. DOX-TPP/HA-ss-OA nanoparticles demonstrate excellent reduction sensitivity, effective tumor cell growth inhibition in vitro, and the ability to overcome drug resistance. Including particle size and zeta potential measurements supports their suitability as drug carriers, highlighting their potential for targeted cancer therapy and further development.

Chemical Profile of Kumquat (Citrus japonica var. margarita) Essential Oil, In Vitro Digestion, and Biological Activity

Kumquat is one of the smallest citrus fruits (from the Rutaceae family), and its essential oil’s biological effects have not yet been sufficiently researched, in contrast to the essential oils of its relatives. Therefore, the aim of this large-scale study was to investigate the chemical profile of kumquat essential oils (KEOs) isolated by microwave-assisted distillation (MAD) and Clevenger hydrodistillation using GC-MS analysis. To test the bioaccessibility of their bioactive components, in vitro digestion with commercially available enzymes was performed. The final step of this research was to test their cytotoxic activity against a cervical cancer cell line (HeLa), a human colon cancer cell line (HCT116), a human osteosarcoma cell line (U2OS), and a healthy cell line (RPE1). Two methods were used to test the antioxidant activity: DPPH (2,2-diphenyl-1-picrylhydrazyl) and ORAC (oxygen radical absorbance capacity). The antibacterial activity was tested in relation to the growth and adhesion of Escherichia coli and Staphylococcus aureus on a polystyrene surface. The GC-MS analysis showed that the major compound in both kumquat essential oils was limonene, which was stable before and after in vitro digestion (>90%). The results showed that the cytotoxic activity of the KEOs in all three cancer cell lines tested was IC50 1–2 mg/mL, and in the healthy cell line (RPE1), the IC50 value was above 4 mg/mL. The antibacterial activity of the KEOs obtained after MAD and Clevenger hydrodistillation was 4 mg/mL against E. coli and 1 mg/mL against S. aureus. The KEOs after MAD and Clevenger hydrodistillation reduced the adhesion of E. coli by more than 1 log, while there was no statistically significant effect on the adhesion of S. aureus to the polystyrene surface. Both KEOs exhibited comparable levels of antioxidant activity using both methods tested, with IC50 values of 855.25 ± 26.02 μg/mL (after MAD) and 929.41 ± 101.57 μg/mL (after Clevenger hydrodistillation) for DPPH activity and 4839.09 ± 91.99 μmol TE/g of EO (after MAD) and 4928.78 ± 275.67 μmol TE/g of EO (after Clevenger hydrodistillation) for ORAC. The results obtained show possible future applications in various fields (e.g., in the food, pharmaceutical, cosmetic, and agricultural industries).

Fibrin Nanoparticles as a Promising Anti-cancer Activity on MCF-7 Cells

Background: Drug delivery methods based on nanoparticles have shown promise in the treatment of cancer. The nanoparticles (NPs) have a cytotoxicity effect on cancer cells. Objectives: This study aimed to evaluate the effect of fibrin nanoparticles (FNPs) on breast cancer cells. Methods: MCF-7 cell lines were treated with 100 µg/mL of FNP for 5 days. On day five, gene and protein expression were evaluated to determine the anti-cancer effects of the FNPs. Results: BCL2, an anti-apoptotic (BCL2) and angiogenic (VEGF) genes and proteins were downregulated in the FNP treatment in comparison with control cells. Conclusions: The study's findings indicate that FNPs considerably reduce anti-apoptotic and angiogenic gene expression in human breast cancer cells.

Sonodynamic Cancer Therapy by Mn(I)-tricarbonyl complexes via Ultrasound-triggered CO release and ROS generation.

A novel ferrocene conjugated Mn(I)-tricarbonyl complex viz [Mn(Fc-tpy)(CO)3Br] (Mn2) where, Fc-tpy = 4'-ferrocenyl-2,2':6',2''-terpyridine was synthesized and fully characterized along with its non-ferrocene analog [Mn(Ph-tpy)(CO)3 Br] Ph-tpy = 4'-phenyl-2,2':6',2''-terpyridine (Mn1) for ultrasound (US) activated anticancer applications. The X-ray structure of Mn2 confirmed its distorted octahedral geometry. Mn1 and Mn2, for the first time, showed US-triggered release of CO and ROS generation (1O2 and •OH) in an aqueous solution from any Mn(I)-tricarbonyl complexes, indicating its potential for synergetic CO gas therapy and sonodynamic therapy. The above-mentioned in-solution chemistry was successfully translated into in vitro cellular models. These complexes showed unprecedented US-triggered toxicity against T-cell lymphoma and human breast cancer cells (IC50 for Mn2 < 1 μM) while were minimally toxic without US or against normal spleen cells. Mn2 was ca. 12 fold more anticancer active than Mn1, indicating that the ferrocene conjugation augmented the US sensitivity. The apoptotic sonotoxicity of Mn2 was due to US-promoted mitochondrial depolarization via ROS generation and CO release. The apoptosis was triggered by caspase 3 activation. This is the first report of Mn(I)-tricarbonyl-based sonosensitizers for cancer SDT. Overall, this study, for the first time, establishes the effectiveness of 3d metal carbonyls in SDT.

Effect of carbon black and silicon dioxide nanoparticle exposure on corona receptor ACE2 and TMPRSS2 expression in the ocular surface.

The Coronavirus disease 2019 (COVID-19) pandemic has led to a global health crisis, including ocular symptoms, primarily targeting the Angiotensin-Converting Enzyme 2 (ACE2) receptor. PM2.5 air pollution may increase infection risk by altering ACE2 expression. Silicon Dioxide (SiO2) and carbon black (CB), major components of PM2.5 from sands and vehicle emissions, were studied for their effects on ACE2 and Transmembrane Protease Serine 2 (TMPRSS2) expression in corneal and conjunctival cells, and ocular tissues. Human corneal epithelial cells (HCECs) and conjunctival epithelial cells (HCjECs) were exposed to nanoparticles (NPs) for 24 hours, assessing viability via WST-8 assay. TNF-α, IL-6, and IL-1β levels in the medium were measured. An in vivo rat study administered NPs via eye drops, with Rose Bengal staining to evaluate damage. ACE2, TMPRSS2, and Angiotensin II (AngII) protein expressions were quantified by Western blot. ACE2 expression in HCjECs increased with NP exposure, while it decreased in HCECs. CB exposure increased TNF-α, IL-6, and IL-1β levels in HCECs. In vivo, corneal exposure to CB decreased ACE2 expression, whereas conjunctival exposure to SiO2 increased ACE2 expression. These changes suggest that SiO2 exposure may increase the risk of COVID-19 through the ocular surface, while CB exposure may decrease it.

M6A-methylase METTL3 promotes retinal angiogenesis through modulation of metabolic reprogramming in RPE cells.

Retinal neovascularization (RNV) disease is one of the leading causes of blindness, yet the molecular underpinnings of this condition are not well understood. To delve into the critical aspects of cell-mediated angiogenesis, we analyzed our previously published single-cell data. Our analysis revealed that retinal pigment epithelium (RPE) cells serve a crucial promotional function in angiogenesis. RPE cells were regulated by N6-methyladenosine (m6A). Next, we detected several critical m6A methylase in hypoxic ARPE-19 cells and in oxygen-induced retinopathy (OIR) mice, our results revealed a significant decrease in the level of methyltransferase like 3 (METTL3). METTL3 specific inhibitor STM2457 intravitreal injection or METTL3 conditional knockout mice both showed a significantly reduced neovascularization area of retina. Additionally, the angiogenesis-related abilities of human retinal endothelial cells (HRECs) were diminished after co-cultured with ARPE-19 treated with STM2457 or sh-METTL3 in vitro. Furthermore, through the integration of Methylated RNA immunoprecipitation (MeRIP) sequencing and RNA sequencing, we discovered that the metabolic enzyme quinolinate phosphoribosyltransferase (QPRT) was directly modified by METTL3 and recognized by the YTH N6-methyladenosine RNA binding protein C1 (YTHDC1). Moreover, after over-expressing QPRT, the angiogenic abilities of HRECs were improved through the phosphorylated phosphatidylinositol-3-kinase (p-PI3K)/ phosphorylated threonine kinase (p-AKT) pathway. Collectively, our study provided a novel therapeutic target for retinal angiogenesis.

Single-shot quantitative differential phase contrast microscope using a single calcite beam displacer

This paper presents the development of a single-shot, partially spatially coherent quantitative differential phase contrast microscopy (Q-DPCM) system. The optical scheme comprises a polarizer, lenses, calcite beam displacer, and analyzer, which can be seamlessly integrated to an existing bright-field microscopy system, transforming it into a Q-DPCM system. It utilizes a partially spatially coherent light source, enabling single-shot quantitative differential phase recovery of the specimens with high spatial phase sensitivity. It generates highly sensitive quantitative differential phase images of the specimens along one direction, like a gradient light interference microscopy (GLIM) system, using only a single interferogram. First, we validated the differential phase measurement capability of the system through experiments on polystyrene spheres (diameter 5.2 µm) and HeLa cells. Next, the system is utilized to generate quantitative phase maps of human red blood cells using two orthogonal differential interferograms recorded at two orientations of the calcite beam displacer. Further, the Q-DPCM system is implemented for 1-h time-lapse live cell monitoring, revealing the dynamics of intracellular granules such as nucleolus and lipids in U2OS cells.

METTL3 accelerates staphylococcal protein A (SpA)-induced osteomyelitis progression by regulating m6A methylation-modified miR-320a.

Osteomyelitis (OM) is an inflammatory disease of bone infection and destruction characterized by dysregulation of bone homeostasis. Staphylococcus aureus (SA) has been reported to be the most common pathogen causing infectious OM. Recent studies have demonstrated that N6-methyladenosine (m6A) regulators are associated with the development of OM. However, the molecular mechanism of m6A modifications in OM remains unclear. Here, we investigated the function of methyltransferase-like 3 (METTL3)-mediated m6A modification in OM development. In this study, human bone mesenchymal stem cells (hBMSCs) were treated with staphylococcal protein A (SpA), a vital virulence factor of SA, to construct cell models of OM. Firstly, we found that METTL3 was upregulated in OM patients and SpA-induced hBMSCs, and SpA treatment suppressed osteogenic differentiation and induced oxidative stress and inflammatory injury in hBMSCs. Functional experiments showed that METTL3 knockdown alleviated the inhibition of osteogenic differentiation and the promotion of oxidative stress and inflammation in SpA-treated hBMSCs. Furthermore, METTL3-mediated m6A modification upregulated miR-320a expression by promoting pri-miR-320a maturation, and the mitigating effects of METTL3 knockdown on SpA-mediated osteogenic differentiation, oxidative stress and inflammatory responses can be reversed by miR-320 mimic. In addition, we demonstrated that phosphatidylinositol-4, 5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) was a downstream target of miR-320a, upregulation of PIK3CA alleviated miR-320a-induced inhibition of osteogenic differentiation, and upregulation of oxidative stress and inflammatory responses during SpA infection. Finally, we found that silencing METTL3 alleviated OM development by regulating the miR-320a/PIK3CA axis. Taken together, our data demonstrated that the METTL3/m6A/miR-320a/PIK3CA axis regulated SpA-mediated osteogenic differentiation, oxidative stress, and inflammatory responses in OM, which may provide a new therapeutic strategy for OM patients.

NEK2 Promotes ESCC Malignant Progression by Inhibiting Cellular Senescence via the FOXM1/c-Myc/p27 Signaling Pathway.

Never in mitosis gene A (NIMA)-related kinase 2 (NEK2) is a crucial serine-threonine kinase involved in the process of cell mitosis. However, the precise relationship between NEK2 and esophageal squamous cell carcinoma (ESCC) remains inadequately understood. NEK2 expression in ESCC tissues was assessed through bioinformatics analysis, reverse transcription-quantitative PCR (RT-qPCR) and immunohistochemistry, revealing a correlation with ESCC patient prognosis. Cultured ESCC cells and human normal esophageal epithelial cells (HEEC) were used to investigate the effects of NEK2 knockdown on the development and progression of ESCC by integrated confluence algorithm, colony formation, wound-healing, transwell, and ESCC xenograft tumor model, in vitro and in vivo. In ESCC tissues, NEK2 was found to be significantly upregulated, and its expression correlated with poor prognosis in ESCC patients. NEK2 may facilitate ESCC development by regulating cell proliferation, migration, and invasion. Additionally, results from in vivo experiments suggested that NEK2 knockdown can inhibit tumor growth. Moreover, forkhead box M1 (FOXM1) was identified as a potential downstream target of NEK2 in the regulation of ESCC, with its overexpression reversing the effects of NEK2 knockdown on ESCC. Mechanistic studies also indicated that NEK2 may promote the malignant progression of ESCC by inhibiting cellular senescence through the activation of the FOXM1/c-Myc/p27 signaling pathways, which may provide a novel perspective for the management of ESCC.

Phenolic Compound Profiles, Antioxidant, and Cytoprotective Activity of Elaeagnus conferta Roxb. Fruit.

In this paper, three varieties of Elaeagnus conferta Roxb fruits prepared by ultrasonic-assisted extraction from a subtropical region southwest of China were utilized as raw materials to investigate their phenolic profiles, antioxidant activities, and protective effects on injured human umbilical vein endothelial cells (HUVECs). The ultra performance liquid chromatography-electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) findings revealed that fifteen substances, including seven phenolic acids, seven flavonoids, and one gallic acid derivative, were discovered. The dihydromyricetin, ellagic acid, gallic acid were the predominant phenolic compounds in all E.conferta fruits. These E.conferta fruits extracts shown excellent antioxidant activity varied from 2.258±0.03~7.844±0.39 μM Trolox/g and protective effect on HUVECs injured by H2O2 through decrease the level of ROS, MDA, LDH and enhance the SOD level. These finding indicate that E.conferta is a valuable source of high-capacity antioxidants that might be used as an alternative material for food industries.

Stochastically structured illumination microscopy scan less super resolution imaging

AbstractIn super-resolution, a varying illumination image stack is required. This enriched dataset typically necessitates precise mechanical control and micron-scale optical alignment and repeatability. Here, we introduce a novel methodology for super-resolution microscopy called stochastically structured illumination microscopy (S2IM), which bypasses the need for illumination control exploiting instead the random, uncontrolled movement of the target object. We tested our methodology within the clinically relevant ophthalmoscopic setting, harnessing the inherent saccadic motion of the eye to induce stochastic displacement of the illumination pattern on the retina. We opted to avoid human subjects by utilizing a phantom eye model featuring a retina composed of human induced pluripotent stem cells (iPSC) retinal neurons and replicating the ocular saccadic movements by custom actuators. Our findings demonstrate that S2IM unlocks scan-less super-resolution with a resolution enhancement of 1.91, with promising prospects also beyond ophthalmoscopy applications such as active matter or atmospheric/astronomical observation.

Suppression of CDK1/Drp1-Mediated Mitochondrial Fission Attenuates Dexamethasone-Induced Extracellular Matrix Deposition in the Trabecular Meshwork.

Aims: Deposition of extracellular matrix (ECM) in the trabecular meshwork (TM), as induced by dexamethasone (Dex), is believed to play an important role in the onset of glucocorticoid-induced glaucoma (GIG). Abnormal ECM deposition is a consequence of mitochondrial dysfunction. We aimed to clarify how mitochondrial dysfunction leads to ECM deposition within the TM and to support the development of novel therapeutic strategies. Results: In primary human TM cells (pHTMCs) and a Dex acetate-induced murine model of GIG, glucocorticoid administration stimulated both mitochondrial fission and ECM deposition. Excessive mitochondrial fission leads to dysfunction and the overexpression of ECM proteins in pHTMCs. Notably, when pHTMCs were treated with the dynamin-related protein 1 (Drp1) inhibitor Mdivi-1 or with Drp1 siRNA, we observed a marked reduction in Dex-induced mitochondrial damage and ECM proteins in vitro. Furthermore, in C57BL/6J mice, treatment with Mdivi-1 mitigated mitochondrial damage and blocked ECM deposition within the TM. We then used Ro3306 to inhibit the cyclin-dependent kinase (CDK)1-mediated phosphorylation of Drp1 at Ser 616, which restored mitochondrial function and diminished Dex-induced ECM protein expression in pHTMCs. Innovation: This study illuminates the pathogenic mechanism linking mitochondrial dysfunction to ECM deposition in GIG. Our innovative approach revealed that Dex stimulates mitochondrial fission via CDK1-mediated p-Drp1s616 overexpression, which drives ECM accumulation. It offered a novel therapeutic strategy for reducing ECM protein expression by inhibiting excessive mitochondrial fission and restoring mitochondrial function. Conclusion: By targeting the CDK1/Drp1-driven mitochondrial fission process, we can counteract Dex-induced ECM deposition in the TM both invivo and in vitro.