Cell Apoptotic Rate Research Articles

Procyanidin B2 alleviates oxidized low-density lipoprotein-induced cell injury, inflammation, monocyte chemotaxis, and oxidative stress by inhibiting the nuclear factor kappa-B pathway in human umbilical vein endothelial cells

BackgroundOxidized low-density lipoprotein (ox-LDL) can initiate and affect almost all atherosclerotic events including endothelial dysfunction. In this text, the role and underlying molecular basis of procyanidin B2 (PCB2) with potential anti-oxidant and anti-inflammatory activities in ox-LDL-induced HUVEC injury were examined.MethodsHUVECs were treated with ox-LDL in the presence or absence of PCB2. Cell viability and apoptotic rate were examined by CCK-8 assay and flow cytometry, respectively. The mRNA and protein levels of genes were tested by RT-qPCR and western blot assays, respectively. Potential downstream targets and pathways of apple procyanidin oligomers were examined by bioinformatics analysis for the GSE9647 dataset. The effect of PCB2 on THP-1 cell migration was examined by recruitment assay. The effect of PCB2 on oxidative stress was assessed by reactive oxygen species (ROS) level, malondialdehyde (MDA) content, and mitochondrial membrane potential (MMP).Resultsox-LDL reduced cell viability, induced cell apoptosis, and facilitated the expression of oxidized low-density lipoprotein receptor 1 (LOX-1), C-C motif chemokine ligand 2 (MCP-1), vascular cell adhesion protein 1 (VCAM-1) in HUVECs. PCB2 alleviated ox-LDL-induced cell injury in HUVECs. Apple procyanidin oligomers triggered the differential expression of 592 genes in HUVECs (|log2fold-change| > 0.58 and adjusted p-value < 0.05). These dysregulated genes might be implicated in apoptosis, endothelial cell proliferation, inflammation, and monocyte chemotaxis. PCB2 inhibited C-X-C motif chemokine ligand 1/8 (CXCL1/8) expression and THP-1 cell recruitment in ox-LDL-stimulated HUVECs. PCB2 inhibited ox-LDL-induced oxidative stress and nuclear factor kappa-B (NF-κB) activation in HUVECs.ConclusionPCB2 weakened ox-LDL-induced cell injury, inflammation, monocyte recruitment, and oxidative stress by inhibiting the NF-κB pathway in HUVECs.

Abstract 5452: AXIN2 differentially modulates stem cell subpopulations providing a potential therapeutic opportunity for targeting colorectal cancer stem cells

Abstract Constitutively activated WNT-signaling promotes stem cell (SC) overpopulation which drives most colorectal cancer (CRC) development. It is not known, however, how WNT-signaling activity regulates different SC sub-populations. We previously reported that SC population size decreases as AXIN2 is knocked down, and increases when AXIN2 is overexpressed. Changes in cell cycle and apoptotic rate were also observed. Here, we present our recent results on changes between different SC subpopulations (ALDH+ vs. LGR5+) that occur due to AXIN2 modulation. We hypothesize that modulation of AXIN2 promotes differential expansion between different CRC SC subpopulations (ALDH+ and LGR5+). We observed that AXIN2 not only increases LGR5+ SC population size, but also expands the ALDH+ SC population to a greater extent as shown by an increased ALDH+-to-LGR5+ SCs ratio. This indicates that AXIN2, via WNT signaling, regulates SC population size, particularly ALDH+ SCs. Since the ALDH SC marker is also a key component in the retinoic acid (RA) signaling pathway, we surmised that increased AXIN2 would not only lead to elevated ALDH levels, but also upregulate the RA-signaling pathway components in CRC cells. To this end, we conducted NanoString mRNA profiling to study the effects of AXIN2 knockdown and overexpression in HT29 and SW480 CRC cells. The results showed that increased AXIN2 significantly up-regulated several key components of the RA-signaling pathway. Taken together, our results indicate that AXIN2 drives a shift in SC subpopulation sizes favoring the ALDH+ SCs. It also promotes the RA-signaling component expression - a pathway known to regulate SC differentiation. As a potential treatment strategy for CRC, therapeutically inducing high levels of AXIN2 could shift the balance between SC populations towards ALDH+ SCs, which may provide a way to target cancer SCs with retinoids to induce their differentiation. Citation Format: Chi Zhang, Caroline O. Facey, Lynn M. Opdenaker, Bruce M. Boman. AXIN2 differentially modulates stem cell subpopulations providing a potential therapeutic opportunity for targeting colorectal cancer stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5452.

Mathematical modeling and dynamic analysis for cancer resistance incorporating persister cells

Drug resistance is a key impediment to cancer treatment, however, the resistance mechanism remains controversial. Experiment evidence indicated that persister cells, a subpopulation in a transient pseudo-dormant state, are posited to play a potential role in the emergence of resistance. In this study, we propose a novel mathematical model for describing the interactions among sensitive, persister, and resistant cells to qualitatively and quantitatively analyze how persister cells affect cancer evolution and unravel the underlying resistance mechanisms. The proposed model is validated by theoretical analysis and fitting of actual human CT scan and mice data. Theoretical analysis demonstrates the existence and stability of the multiple steady states, as well as the global asymptotical stability of a unique steady state. Furthermore, bifurcation analysis reveals that the key factors, including the apoptotic rate of sensitive cells and the transformation rate between sensitive and persister cells, induce bistable phenomenon and are characterized by double saddle–node and transcritical bifurcations. The bistable region indicates that persister cells under high stable steady state as a reservoir could promote the emergence of resistant cells conferring a proliferative advantage and conversely when they are in lower stable steady state, the source of resistant cells and resistance process will be controlled. Therefore, the first transcritical bifurcation point can be viewed as an early indicator to detect critical transitions from low resistance to high resistance. Together, the proposed model and quantitative results would provide new insights for searching for strategies in modulating and decreasing the risk of drug resistance.

CircMCTP2 enhances the progression of bladder cancer by regulating the miR-99a-5p/FZD8 axis

BackgroundCircRNAs and miRNAs are involved in the progression of tumor. CircMCTP2 is considered as a novel tumor promoter. However, the exact functions of circMCTP2 in bladder cancer are still unclear. This study was designed to explore the underlying mechanisms of circMCTP2-modulated tumor development in bladder cancer.MethodsThe present study is an original research. The levels of circMCTP2 in a total of 39 bladder cancer specimens and cell lines were determined by RT-qPCR. The expression of FZD8 in T24 and RT-4 cells treated with miR-99a-5p mimics were examined using western blotting. In addition, the proliferative, migrative and invasive abilities of transfected cells were determined by CCK8 and Transwell assays. Furthermore, the apoptosis of transfected cells was evaluated using flow cytometry. Dual luciferase reporter assay was performed to elucidate the relationship between miR-99a-5p and circMCTP2/FZD8.ResultsThe levels of circMCTP2 were elevated in bladder cancer samples and cells, and this was related to worse survival rate. Downregulation of circMCTP2 suppressed growth and metastasis of cells, whereas the apoptotic rate of cells was enhanced. The levels of miR-99a-5rp was elevated after the downregulation of circMCTP2. Moreover, reverse correlation between the expression of miR-99a-5p and circMCTP2 was revealed in bladder cancer specimens. Additionally, FZD8 was the putative target of miR-99a-5p and the mimics of miR-99a-5p inhibited the proliferation, migration and invasion of bladder cancer cells via the FZD8/Wnt-b-catenin axis. Moreover, circMCTP2 regulated the growth and metastasis of bladder cancer cells potentially through regulating the miR-99a-5p/FZD8/Wnt-b-catenin axis. In summary, circMCTP2 was considered as an oncogenic factor through regulating the miR-99a-5p/FZD8/Wnt-b-catenin axis.ConclusionsThis novel signaling could regulate the biological behaviours of bladder cancer cells, and these findings highlighted circMCTP2 as a critical target for treating bladder cancer.

C/EBPα-p30 confers AML cell susceptibility to the terminal unfolded protein response and resistance to Venetoclax by activating DDIT3 transcription

BackgroundAcute myeloid leukemia (AML) with biallelic (CEBPAbi) as well as single mutations located in the bZIP region is associated with a favorable prognosis, but the underlying mechanisms are still unclear. Here, we propose that two isoforms of C/EBPα regulate DNA damage-inducible transcript 3 (DDIT3) transcription in AML cells corporately, leading to altered susceptibility to endoplasmic reticulum (ER) stress and related drugs.MethodsHuman AML cell lines and murine myeloid precursor cell line 32Dcl3 cells were infected with recombinant lentiviruses to knock down CEBPA expression or over-express the two isoforms of C/EBPα. Quantitative real-time PCR and western immunoblotting were employed to determine gene expression levels. Cell apoptosis rates were assessed by flow cytometry. CFU assays were utilized to evaluate the differentiation potential of 32Dcl3 cells. Luciferase reporter analysis, ChIP-seq and ChIP-qPCR were used to validate the transcriptional regulatory ability and affinity of each C/EBPα isoform to specific sites at DDIT3 promoter. Finally, an AML xenograft model was generated to evaluate the in vivo therapeutic effect of agents.ResultsWe found a negative correlation between CEBPA expression and DDIT3 levels in AML cells. After knockdown of CEBPA, DDIT3 expression was upregulated, resulting in increased apoptotic rate of AML cells induced by ER stress. Cebpa knockdown in mouse 32Dcl3 cells also led to impaired cell viability due to upregulation of Ddit3, thereby preventing leukemogenesis since their differentiation was blocked. Then we discovered that the two isoforms of C/EBPα regulate DDIT3 transcription in the opposite way. C/EBPα-p30 upregulated DDIT3 transcription when C/EBPα-p42 downregulated it instead. Both isoforms directly bound to the promoter region of DDIT3. However, C/EBPα-p30 has a unique binding site with stronger affinity than C/EBPα-p42. These findings indicated that balance of two isoforms of C/EBPα maintains protein homeostasis and surveil leukemia, and at least partially explained why AML cells with disrupted C/EBPα-p42 and/or overexpressed C/EBPα-p30 exhibit better response to chemotherapy stress. Additionally, we found that a low C/EBPα p42/p30 ratio induces resistance in AML cells to the BCL2 inhibitor venetoclax since BCL2 is a major target of DDIT3. This resistance can be overcome by combining ER stress inducers, such as tunicamycin and sorafenib in vitro and in vivo.ConclusionOur results indicate that AML patients with a low C/EBPα p42/p30 ratio (e.g., CEBPAbi) may not benefit from monotherapy with BCL2 inhibitors. However, this issue can be resolved by combining ER stress inducers.

Bisphenol A promotes cell death in healthy respiratory system cells through inhibition of cell proliferation and induction of G2/M cell cycle arrest.

Bisphenol A (BPA) is a substance that can harm the environment and human health by interfering with the normal functioning of the body's hormonal system. It is commonly found in various plastic-based products such as cosmetics, canned foods, beverage containers, and medical equipment and as well as it can also be absorbed by inhalation. There have been limited studies on the effects of BPA on lung fibroblasts, and it is still unclear how high levels of BPA can impact respiratory system cells, particularly the lungs and trachea. In this research, we aimed to investigate the cell cycle disruption potential of BPA on respiratory system cells by examining healthy trachea and lung cells together for the first time. The findings indicated that BPA exposure can alter the healthy cells' morphology, leading to reduced cellular viability that has been assessed by MTT and SRB assays. BPA treatment was able to activate caspase3 as expected, which could cause apoptosis in treated cells. Although the highest dose of BPA did not increase the apoptotic rate of rat trachea cells, it remarkably caused them to become necrotic (52.12%). In addition to quantifying the induction of apoptosis and necrosis by BPA, cell cycle profiles were also determined using flow cytometry. Thereby, BPA treatment unexpectedly inhibited the cell cycle's progression by causing G2/M cell cycle arrest in both lung and tracheal cells, which hindered cell proliferation. The findings of the study suggested that exposure to BPA could lead to serious respiratory problems, even respiratory tract cancers via alterations in the cell cycle.

USP48 deubiquitination stabilizes SLC1A5 to inhibit retinal pigment epithelium cell inflammation, oxidative stress and ferroptosis in the progression of diabetic retinopathy.

Diabetic retinopathy is one of the complications of diabetes mellitus. The aim of this study was to explore the effects of ubiquitin-specific protease 48 (USP48) and its underlying mechanisms in the development of diabetic retinopathy. CCK-8 assay, EdU assay, and flow cytometry were used to measure the proliferative ability and the apoptotic rate of ARPE-19 cells, respectively. ELISA kits were utilized to assess the levels of inflammatory cytokines. The levels of Fe2+, ROS and MDA were detected using the corresponding biochemical kits. The protein expression of USP48 and SLC1A5 was examined through western blot. The mRNA level of SLC1A5 was determined using RT-qPCR. The interaction relationship between USP48 and SLC1A5 was evaluated using Co-IP assay. High glucose (HG) treatment significantly inhibited cell proliferation and elevated cell apoptosis, inflammation, ferroptosis and oxidative stress in ARPE-19 cells. HG treatment-caused cell damage was hindered by USP48 or SLC1A5 overexpression in ARPE-19 cells. Fer-1 treatment improved HG-caused cell damage in ARPE-19 cells, which was blocked by USP48 knockdown. Moreover, USP48 knockdown decreased SLC1A5 expression. SLC1A5 downregulation reversed the improvement effects of USP48 upregulation on cell damage in HG-treated ARPE-19 cells. USP48 overexpression deubiquitinated SLC1A5 to elevate cell proliferation and suppress cell apoptosis, inflammation, ferroptosis and oxidative stress in HG-triggered ARPE-19 cells, thereby inhibiting the progression of diabetic retinopathy.

Rho kinase inhibitor Y27632 promotes survival of human induced pluripotent stem cells during differentiation into functional midbrain dopaminergic progenitor cells in vitro

To improve the efficiency of induced differentiation of primitive neural epithelial cells derived from human induced pluripotent stem cells (hiPSCs-NECs) into functional midbrain dopaminergic progenitor cells (DAPs). HiPSCs were cultured in mTeSRTM medium containing DMH1 (10 μmol/L), SB431542 (10 μmol/L), SHH (200 ng/mL), FGF8 (100 ng/mL), purmorphamine (2 μmol/L), CHIR99021 (3 μmol/L), and N2 (1%) for 12 days to induce their differentiation into primitive neuroepithelial cells (NECs). The hiPSCs-NECs were digested with collagenase Ⅳ and then cultured in neurobasal medium supplemented with 1% N2, 2% B27-A, BDNF (10 ng/mL), GDNF (10 ng/mL), AA, TGF-β, cAMP, and 1% GlutaMax in the presence of different concentrations of Rho kinase inhibitor Y27632, and the culture medium was changed the next day to remove Y27632. Continuous induction was performed until day 28 to obtain DAPs. Human iPSCs expressed the pluripotency markers OCT4, SOX2, Nanog, and SSEA1 and were positive for alkaline phosphatase staining. The hiPSCs-NECs were obtained on day 13 in the form of neural rosettes expressing neuroepithelial markers SOX2, nestin, and PAX6. In digested hiPSCs-NECs, the addition of 5 μmol/L Y27632 significantly promoted survival of the adherent cells, increased cell viability and the proportion of S-phase cells (P < 0.01), and reduced the rate of apoptotic cells (P < 0.05). On day 28 of induction, the obtained cells highly expressed the specific markers of DAPS (TH, FOXA2, NURR1, and Tuj1). Treatment with Y27632 (5 μmol/L) for 24 h significantly promotes the survival of human iPSCs-NECs during their differentiation into DPAs without affecting the cell differentiation, which indirectly enhances the efficiency of cell differentiation.

The safety evaluation of Shenze Shugan capsule and mechanism of apoptosis induced by five potentially nephrotoxic components

Ethnopharmacological relevanceShenze Shugan capsule is a prescription of traditional Chinese medicine for nonalcoholic steatohepatitis treatment. It includes Rhei Radix et Rhizoma (RR), Cassiae Semen (CS) and Alismatis Rhizoma(AR), which widely contains rhein, emodin, aurantio-obtusin, alisol A and alisol B 23-monoacetate. Aim of the studyIn this study, we aimed to explore the safety of the medicine, and further elucidate the mechanism of apoptosis induction in HK-2 cells by five components, including rhein, emodin, aurantio-obtusin, alisol A and alisol B 23-monoacetate. Materials and methodsWe investigated the nephrotoxicity of Shenze Shugan capsule, including RR, CS, AR and mixed herbs given for two months in rats. Superoxide dismutase (SOD) in kidney tissues, urea nitrogen (BUN) and creatinine (CRE) in serum were detected, and renal pathology analysis was performed. In cell experiments, the apoptotic rate and cell cycle distribution of HK-2 cells were tested by flow cytometry. The levels of mitochondrial membrane potential (ΔΨm) and related protein expression in mitochondrial pathway were measured as well. ResultsWe confirmed that two months of administering high doses(60 times the dose for clinical use in adults) of RR, CS or mixed herbs upregulated the levels of CRE and RUN, inhibited SOD activity, and increased the degree of tubular degeneration and glomerular dilatation, but Shenze Shugan capsule has no significant differences in renal structure or renal function. In addition, we found that five components all concentration-dependently inhibited HK-2 cells proliferation and induced apoptosis, especially aurantio-obtusin as the novel nephrotoxic component. Rhein and emodin significantly induced S/M accumulation, but aurantio-obtusin, alisol A and alisol B 23-monoacetate significantly induced G1/M accumulation in HK-2 cells. Similarly, they could induce Caspase3 activation, loss of mitochondrial membrane potential (ΔΨm), and down-regulation of Bcl-2 and up-regulation of Bax. ConclusionsThrough a two-month subchronic toxicity study in rats, our preliminary determination is that this formulation is safe and reliable for long-term use. Interestingly, the potentially toxic herbs such as RR, CS, AR can reduce toxicity by drug compatibility. When further exploring the mechanism of action of toxic herbs, we found that mitochondrial pathway is involved in the apoptosis of HK -2 cells induced by rhein, emodin, aurantio-obtusin, alisol A and alisol B 23-monoacetate. Our findings provide new ideas for safety studies of Shenze Shugan capsule.

Synthesis and biological evaluations of 8-biaryl-2,2-dimethylbenzopyranamide derivatives against Alzheimer's disease and ischemic stroke

Alzheimer's disease, the commonest cause of dementia, is a growing global health concern with huge implications for individuals and society. Stroke has still been a significant challenge in clinics for a long time, which is the second leading cause of death in the world, especially ischemic stroke. Both Alzheimer's disease and stroke are closely related to oxidative stress and HIF-1 signaling pathways in nerve cells. Herein, we describe our structure-based design, synthesis, and biological evaluation of a new class of 8-biaryl-2,2-dimethylbenzopyranamide derivatives as natural product derivatives. Our efforts have resulted in the discovery of highly potent neuroprotective agents, as exemplified by compound D13 as a HIF-1α inhibitor, which significant improvement in the behavior of Alzheimer's disease mice and shows great potential improvement of brain infarct volume in pMCAO model rats, improves the increase of blood–brain barrier permeability after cerebral ischemia in rats, neuroprotective effect, reduce the level of apoptotic cells in rats after cerebral ischemia, better than Edaravone.

Killing effect of anti-MSLN-iCAR-NK cells derived from induced pluripotent stem cells on ovarian epithelial cancer cells

Objective: To investigate the cytotoxic effects of induced pluripotent stem (iPS) cells of anti-mesothelin (MSLN)-chimeric antigen receptor natural killer (CAR-NK) cells (anti-MSLN-iCAR-NK cells) on ovarian epithelial cancer cells. Methods: Twenty cases of ovarian cancer patients who underwent surgical treatment at Henan Provincial People's Hospital from September 2020 to September 2021 were collected, and 20 cases of normal ovarian tissues resected during the same period due to other benign diseases were also collected. (1) Immunohistochemistry and immunofluorescence were used to verify the expression of MSLN protein in ovarian cancer tissues. (2) Fresh ovarian cancer tissues were extracted and cultured to obtain primary ovarian cancer cells. Recombinant lentiviral vectors targeting anti-MSLN-CAR-CD244 were constructed and co-cultured with iPS cells to obtain anti-MSLN-iCAR cells. These cells were differentiated into anti-MSLN-iCAR-NK cells using cytokine-induced differentiation method. The cell experiments were divided into three groups: anti-MSLN-iCAR-NK cell group, natural killer (NK) cell group, and control group. (3) Flow cytometry and live cell staining experiment were used to detect the apoptosis of ovarian cancer cells in the three groups. (4) Enzyme-linked immunosorbent assay (ELISA) was used to measure the expression levels of interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), granzyme B (GZMB), perforin 1 (PRF1), interleukin (IL)-6, and IL-10 in the three groups of ovarian cancer cells. Results: (1) Immunohistochemistry analysis showed that a positive expression rate of MSLN protein in ovarian cancer tissues of 65% (13/20), while normal ovarian tissues had a positive rate of 30% (6/20). The comparison between the two groups was statistically significant (χ2=4.912, P=0.027). Immunofluorescence analysis revealed that the positive expression rate of MSLN protein in ovarian cancer tissues was 70% (14/20), while normal ovarian tissues had a positive rate of 30% (6/20). The comparison between the two groups was statistically significant (χ2=6.400, P=0.011). (2) Flow cytometry analysis showed that the apoptotic rate of ovarian cancer cells in the anti-MSLN-iCAR-NK cell group was (29.27±0.85)%, while in the NK cell group and control group were (8.44±0.34)% and (6.83±0.26)% respectively. There were statistically significant differences in the comparisons between the three groups (all P<0.01). Live cell staining experiment showed that the ratio of dead cells to live cells in the anti-MSLN-iCAR-NK cell group was (36.3±8.3)%, while in the NK cell group and control group were (5.4±1.4)% and (2.0±1.3)% respectively. There were statistically significant differences in the comparisons between the three groups (all P<0.001). (3) ELISA analysis revealed that the expression levels of IFN-γ, TNF-α, GZMB, PRF1, IL-6, and IL-10 in ovarian cancer cells of the anti-MSLN-iCAR-NK cell group were significantly higher than those in the NK cell group and the control group (all P<0.05). Conclusion: The anti-MSLN-iCAR-NK cells exhibit a strong killing ability against ovarian cancer cells, indicating their potential as a novel immunotherapy approach for ovarian cancer.

Recognition of Phosphatidylserine Externalisation on Human Chronic Myeloid Leukaemia (K562) Cells Induced by Chrysanthemum morifolium Methanol Extract

Chrysanthemum morifolium is one of the common plants used in traditional Chinese herbal medicine and currently being discovered as an invaluable source for treatment of cancerous diseases. Conventional approach to leukaemia treatment such as chemotherapeutic agents results in good overall survival (OS) in patients but issues of multi-drug resistance (MDR) and disease heterogeneity can make chemotherapy less effective. Thus there is a constant need for alternative therapeutic strategy such as developing new anti-cancer drug from natural products. The present study is carried out mainly to detect phosphatidylserine externalisation as an apoptosis indicator on human chronic myeloid leukaemia (K562) cells induced by Chrysanthemum morifolium methanol extract. K562 cells were treated with indicated IC50 values of bud and flower methanol crude extracts for 24 hours. Phase contrast microscope was used to observe morphological changes in K562 cells post-treatment. Annexin V-FITC/PI apoptosis assay via flow cytometry was done to investigate apoptosis phenomena by detecting phosphatidylserine (PS) externalisation in treated K562 cells. Morphological observations using phase contrast microscopy showed distinct apoptotic characteristics in K562 cells treated with both bud and flower methanol extracts. Annexin V-FITC/PI assay demonstrated a significantly high apoptotic cell population and apoptotic rate (p &lt;0.05) in treated K562 cells compared to control cells. The study outcomes presented promising anti-cancer ability for Chrysanthemum morifolium.

Bioactive Properties of Tagetes erecta Edible Flowers: Polyphenol and Antioxidant Characterization and Therapeutic Activity against Ovarian Tumoral Cells and Caenorhabditis elegans Tauopathy.

Tagetes erecta is an edible flower deeply rooted in traditional Mexican culture. It holds a central role in the most popular and iconic Mexican celebration, "the Day of the Dead". Furthermore, it is currently receiving interest as a potential therapeutic agent, motivated mainly by its polyphenol content. The present study aims to evaluate the biological activity of an extract synthesized from the petals of the edible flower T. erecta. This extract showed significant antioxidant scores measured by the most common in vitro methodologies (FRAP, ABTS, and DPPH), with values of 1475.3 μM trolox/g extr, 1950.3 μM trolox/g extr, and 977.7 μM trolox/g extr, respectively. In addition, up to 36 individual polyphenols were identified by chromatography. Regarding the biomedical aspects of the petal extract, it exhibited antitumoral activity against ovarian carcinoma cells evaluated by the MTS assay, revealing a lower value of IC50 compared to other flower extracts. For example, the extract from T. erecta reported an IC50 value half as low as an extract from Rosa × hybrida and six times lower than another extract from Tulbaghia violacea. This antitumoral effect of T. erecta arises from the induction of the apoptotic process; thus, incubating ovarian carcinoma cells with the petal extract increased the rate of apoptotic cells measured by flow cytometry. Moreover, the extract also demonstrated efficacy as a therapeutic agent against tauopathy, a feature of Alzheimer's disease (AD) in the Caenorhabditis elegans experimental model. Treating worms with the experimental extract prevented disfunction in several motility parameters such as wavelength and swimming speed. Furthermore, the T. erecta petal extract prevented the release of Reactive Oxygen Species (ROS), which are associated with the progression of AD. Thus, treatment with the extract resulted in an approximate 20% reduction in ROS production. These findings suggest that these petals could serve as a suitable source of polyphenols for biomedical applications.

Knocking down of lncRNA SBF2-AS1 inhibits proliferation, invasion and induces apoptosis of colorectal cancer through regulating PTEN.

To illustrate the potential function of lncRNA SBF2-AS1 in the progression of colorectal cancer (CRC) and the molecular mechanism. The relative level of SBF2-AS1 in CRC tissues and cell lines was determined by qRT-PCR. Its level in CRC patients with different tumor stages and tumor sizes was examined. After the knockdown of SBF2-AS1, proliferative, invasive abilities and apoptotic rate of CRC cells were evaluated. The correlation between SBF2-AS1 and PTEN was analyzed in CRC tissues. Furthermore, the subcellular distribution of SBF2-AS1 was assessed. Through RIP and ChIP assay, the interaction between SBF2-AS1 and PTEN was identified. Finally, the involvement of PTEN in SBF2-AS1-mediated CRC progression was analyzed. SBF2-AS1 was upregulated in CRC tissues and cell lines. Its level remained higher in CRC with worse tumor stage and larger tumor size. Knockdown of SBF2-AS1 attenuated proliferative, invasive abilities, but induced apoptotic rate of SW480 and DLD1 cells. A negative correlation was identified between expression levels of SBF2-AS1 and PTEN in CRC tissues. PTEN level was negatively regulated by SBF2-AS1. Subcellular distribution analysis indicated that SBF2-AS1 was mainly expressed in the nucleus. Furthermore, the RIP assay proved the binding of SBF2-AS1 to EZH2 and SUZ12. Knockdown of SBF2-AS1 attenuated the recruitment ability of EZH2 to PTEN. Notably, inhibited proliferation by transfection of sh-SBF2-AS1 1# was partially reversed after co-transfection of sh-PTEN. LncRNA SBF2-AS1 is upregulated in CRC. Knocking down of lncRNA SBF2-AS1 inhibits proliferation, and invasion and induces apoptosis of colorectal cancer by interacting with EZH2 to downregulate PTEN level.

Study on the biological properties of SMILE-derived corneal stromal lenticules after long-term cryopreservation in nutrient capsules

PurposeTo investigate the long-term preservation effects of nutrient capsules on the physiological activity, collagen fiber structure and transmittance of corneal stromal lenticules derived from small incision lenticule extraction (SMILE). MethodsA new nutrient capsule was constructed for long-term preservation of SMILE-derived corneal stromal lenticules. The lenticules were randomly divided into 99% anhydrous glycerol, and hydrogel nutrient capsules. After preserving for 1 year at −80 °C, lenticules were compared with fresh lenticules. The optical transmittance, tissue morphology, ultrastructure, cells activity and immunogenicity of the lenticules was detected and compared between different groups. ResultsThe rate of apoptotic cells was significantly higher in the glycerol group compared with the nutrient capsule group (P < 0.0001). More viable cells were present in the lenticules after nutrient capsule preservation compared to the glycerol group (P = 0.0003). The mean transmittance of the lenticules in the glycerol group (50 ± 18%) was significantly lower (P = 0.0008) compared to the control group (75 ± 11%), and the lenticules transmittance of the nutrient capsule group (64 ± 15%) after long-term preservation was not significantly different (P = 0.23) compared to the control group. The structure of HE staining showed that the collagen fibers in the nutrient capsule group were arranged in parallel and neatly, and a few cavitation vesicles were visible inside the tissue. There was no significant difference in the number of lenticular collagen fibers in the nutritional capsule group compared to the fresh lenticule group (P = 0.06). HLA-DR, HLA-ABC, CD45, CD25 and CD69 expression was low in all groups of lenticules after preservation. ConclusionsNutrient capsules can preserve lenticules for a long time and maintain the transmission structure and cells activity of lenticules.

IGF2BP1 facilitates non-small cell lung cancer progression by regulating the KIF2A-mediated Wnt/β-catenin pathway.

Insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs) are crucially implicated in the cancer progression. The current study intends to excavate and clarify the mechanisms of the key IGF2BPs in non-small cell lung cancer (NSCLC). The expression of IGF2BPs and kinesin family member 2A (KIF2A) was examined using immunohistochemistry, real-time quantitative polymerase chain reaction, and western blot in NSCLC tissue samples or cell lines. NSCLC cell viability was examined using a cell counting kit-8 assay. Cell apoptotic rate was assessed using flow cytometry analysis. The migration and invasion of H1299 cells were subject to scratch test and Transwell assays, respectively. Starbase 2.0 was used to detect the downstream factors of the IGF2BP1 protein. The binding of IGF2BP with KIF2A was detected using RNA binding protein immunoprecipitation assays. Ki-67 immunohistochemistry assay and TUNEL assays were applied for the evaluation of proliferation and apoptosis in vivo, respectively. IGF2BP1 was upregulated in NSCLC tissue samples and cells. Functionally, IGF2BP1 overexpression promoted the proliferative ability, migration, and invasiveness of H1299 cells, while inhibiting cell apoptosis in vitro. In vivo studies revealed that overexpression of IGF2BP1 promoted tumor growth of NSCLC. Mechanistically, IGF2BP1 was involved in KIF2A mRNA stabilization. KIF2A exerted the same functions as IGF2BP1 via the Wnt/β-catenin signaling. In conclusion, IGF2BP1 enhances NSCLC malignant progression by stabilizing KIF2A to modulate the Wnt/β-catenin pathway.

Regulation of Interferon-β-Modified Human Umbilical Cord Mesenchymal Stem Cell-Derived Exosomes in Proliferation and Apoptosis of Prostate Cancer Cells

ABSTRACT Prostate cancer (PCa) poses a serious burden to men. Interferon-β (IFN-β) is implicated in cancer cell growth. This study hence explored the regulation of IFN-β-modified human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSC-Exos) in PCa cells. In vitro-cultured hUCMSCs were transfected with pcDNA3.1-IFN-β plasmid or IFN-β siRNA. hUCMSC-Exos were extracted by ultracentrifugation and identified. PCa cells (PC3 and LNCap) were treated with Exos. Cellular internalization of Exos by cells was detected by uptake assay. Cell proliferation, cycle, and apoptosis were evaluated by CCK-8, EdU staining, and flow cytometry. Levels of cell cycle-related proteins (cyclin D/cyclin E) were determined by Western blot. The effect of IFN-β-modified hUCMSC-Exos in vivo was analyzed. IFN-β-modified hUCMSC-Exos (Exooe- IFN −β or Exosi- IFN −β) were successfully isolated. IFN-β was encapsulated in Exos, and PCa cells could uptake Exos. After treating with Exooe- IFN −β, PCa cell proliferation was impeded, the percentage of cells in the G0/G1 phase, cyclin D/cyclin E levels, and cell apoptotic rate were elevated, while cells treated with Exooe- IFN −β exhibited contrary trends. IFN-β-modified hUCMSC-Exos reduced PCa tumor size and weight in vivo. Conjointly, IFN-β-modified hUCMSC-Exos suppress PCa cell proliferation and facilitate apoptosis.

Repeated radon exposure induced epithelial-mesenchymal transition-like transformation via disruption of p53-dependent mitochondrial function.

As a human carcinogen, radon and its progeny are the second most important risk factor for lung cancer after smoking. The tumor suppressor gene, p53, is reported to play an important role in the maintenance of mitochondrial function. In this work, we investigated the association between p53 and p53-responsive signaling pathways and radon-induced carcinogenesis. After repeated radon exposure, the malignant characteristics, cell cycle arrest, cell apoptotic rate, adenosine triphosphate (ATP) content, reactive oxygen species (ROS) level, mitochondrial DNA (mtDNA) copy number as well as indicative biomarkers involved in mitochondrial energy metabolism were evaluated in BEAS-2B cells or BALB-c mouse lung tissue. Radon exposure induced epithelial-mesenchymal transition (EMT)-like transformation in BEAS-2B cells, as indicated by increased cell proliferation and migration. Additional mitochondrial alterations, including decreased ATP content, increased ROS levels, mtDNA copy numbers, cell apoptosis, and G2/M cell cycle arrest were observed. Radon exposure caused an energy generation shift from aerobic respiration to glycolysis as reflected by increased expression of TIGAR and p53R2 proteins and decreased expression of SCO2 protein in BEAS-2B cells, and increased expression of p53, SCO2 and TIGAR proteins in mouse lung tissue, respectively. The effects of p53 deficiency on the prevention of mitochondrial dysfunction suggested a protective role of p53 in radon-induced malignant-like features in BEAS-2B cells. Repeated radon exposure induced EMT-like transformation in BEAS-2B cells via disruption of mitochondrial function. Activation of p53 and p53-responsive signaling pathways in BEAS-2B cells and BALB-c mice may confer a protective mechanism for radon-induced lung injury.

The selective NOX4 inhibitor GLX7013159 decreases blood glucose concentrations and human beta-cell apoptotic rates in diabetic NMRI nu/nu mice transplanted with human islets

NADPH oxidase 4 (NOX4) inhibition has been reported to mitigate diabetes-induced beta-cell dysfunction and improve survival in vitro, as well as counteract high-fat diet-induced glucose intolerance in mice. We investigated the antidiabetic effects of the selective NOX4 inhibitor GLX7013159 in vivo in athymic diabetic mice transplanted with human islets over a period of 4 weeks. The GLX7013159-treated mice achieved lower blood glucose and water consumption throughout the treatment period. Furthermore, GLX7013159 treatment resulted in improved insulin and c-peptide levels, better insulin secretion capacity, as well as in greatly reduced apoptotic rates of the insulin-positive human cells, measured as colocalization of insulin and cleaved caspase-3. We conclude that the antidiabetic effects of NOX4 inhibition by GLX7013159 are observed also during a prolonged study period in vivo and are likely to be due to an improved survival and function of the human beta-cells.

A Reproducible Cartilage Impact Model to Generate Post-Traumatic Osteoarthritis in the Rabbit.

Post-traumatic osteoarthritis (PTOA) is responsible for 12% of all osteoarthritis cases in the United States. PTOA can be initiated by a single traumatic event, such as a high-impact load acting on articular cartilage, or by joint instability, as occurs with anterior cruciate ligament rupture. There are no effective therapeutics to prevent PTOA currently. Developing a reliable animal model of PTOA is necessary to better understand the mechanisms by which cartilage damage proceeds and to investigate novel treatment strategies to alleviate or prevent the progression of PTOA. This protocol describes an open, drop tower-based rabbit femoral condyle impact model to induce cartilage damage. This model delivered peak loads of 579.1 ± 71.1 N, and peak stresses of 81.9 ± 10.1 MPa with a time-to-peak load of 2.4 ± 0.5 ms. Articular cartilage from impacted medial femoral condyles (MFCs) had higher rates of apoptotic cells (p = 0.0058) and possessed higher Osteoarthritis Research Society International (OARSI) scores of 3.38 ± 1.43 compared to the non-impacted contralateral MFCs (0.56 ± 0.42), and other cartilage surfaces of the impacted knee (p < 0.0001). No differences in OARSI scores were detected among the non-impacted articular surfaces (p > 0.05).