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Triple-negative breast cancer (TNBC) is a malignant breast cancer. There is an urgent need for effective drugs to be developed for TNBC. Tubocapsicum anomalum ( T. anomalum ) has been reported to have an anti-tumor effect, and six novel withanolides were isolated from it and designated as TAMEWs. However, its anti-TNBC effect is still unknown. The results of an MTT assay indicated a higher sensitivity of TNBC cells to TAMEWs compared to other cells. TAMEWs induced apoptosis via mitochondrial dysfunction. They caused increased levels of lipid ROS and Fe 2+ , with downregulation of GSH and cystine uptake, and it has been confirmed that TAMEWs induced ferroptosis. Additionally, the results of Western blotting indicate that TAMEWs significantly decrease the expressions of ferroptosis-related proteins. Through further investigation, it was found that the knockdown of the p53 gene resulted in a significant reversal of ferroptosis and the expressions of its associated proteins SLC7A11, ASCT2, and GPX4. In vivo, TAMEWs suppressed TNBC growth with no obvious damage. The IHC results also showed that TAMEWs induced apoptosis and ferroptosis in vivo. Our findings provide the first evidence that TAMEWs suppress TNBC growth through apoptosis and ferroptosis.

LFZ-4-46, that is [2-hydroxy-1-phenyl-1,5,6,10b-tetrahydropyrazolo(5,1-a) isoquinolin-3(2H)-yl](phenyl) methanone, a tetrahydroisoquinoline derivative with a pyrazolidine moiety, was synthetically prepared. The anti-cancer mechanism of the compound has not been clarified yet. In this study, the anticancer effects and potential mechanisms of LFZ-4-46 on human breast and prostate cancer cells were explored. (a) 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazoliumbromide assay was first performed to detect the effects of LFZ-4-46 on the viability of human cancer cells. (b) Comet assay was utilized to evaluate DNA damage. (c) Cell cycle, apoptosis and mitochondrial membrane potential were detected by flow cytometry. (d) The expression of relative proteins was detected by western blotting assay. LFZ-4-46 significantly inhibited the viability of cancer cells in a time- and dose-dependent manner and had no obviously inhibitory effect on the viability of mammary epithelial MCF-10A cells. Mechanistic studies demonstrated that LFZ-4-46-induced cell apoptosis and cycle arrest were mediated by DNA damage. It caused DNA damage through activating γ-H2AX and breaking DNA strands. Further studies showed that mitogen-activated protein kinasess pathway was involved in these activated several key molecular events. Finally, LFZ-4-46 showed a potent antitumor effect in vivo. These results suggest that LFZ-4-46 may be a potential lead compound for the treatment of breast and prostate cancer., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc.)

Valtrate is a principle compound isolated from Valeriana jatamansi Jones , a traditional Chinese folk medicine originally used to treat various nervous disorders. Here, we found that valtrate exhibited significant anti-cancer activity in vitro , especially in human breast cancer cells, while displayed relatively low cytotoxicity to normal human breast epithelial cells (MCF 10A). Valtrate induced cell cycle arrest at G2/M stage and apoptosis in MDA-MB-231 and MCF-7 cells, with reduced expression of p-Akt (Ser 473), cyclin B1 and caspase 8, and increased expression of p21, p-cdc2, cleaved-caspase 3, cleaved-caspase 7 and poly (ADP-ribose) polymerase (PARP). In addition, valtrate inhibited cell migration through down-regulation of MMP-9 and MMP-2 expression. These results demonstrate that valtrate possesses anti-breast cancer activities via cell cycle arrest, apoptosis, and inhibition of cell migration, thus supporting valtrate as a potential antitumor agent.

The biological function of human ovaries declines along with aging. To identify the underlying molecular changes during ovarian aging, pigs were used as model animals. Genome-wide DNA methylation and transcriptome-wide RNA expression analyses were performed via high-throughput sequencing of ovaries from young pigs (180 days, puberty stage of first ovulation) and old pigs (eight years, reproductive exhaustion stage). The results identified 422 different methylation regions between old and young pigs; furthermore, a total of 2,243 mRNAs, 95 microRNAs, 248 long noncoding RNAs (lncRNAs), and 116 circular RNAs (circRNAs) were differentially expressed during both developmental stages. Gene ontology analysis showed that these genes related to different methylation and expression are involved in the ovarian aging cycle. Specifically, these are involved in cell apoptosis, death effector domain binding, embryonic development, reproduction and fertilization process, ovarian cumulus expansion, and the ovulation cycle. Multigroup cooperative control relationships were also assessed, and competing endogenous RNA (ceRNA) networks were constructed in the ovarian aging cycle. These data will help to clarify ovary age-associated potential molecular changes in DNA methylation and transcriptional patterns over time. [ABSTRACT FROM AUTHOR]

Accumulating evidence has indicated that microRNAs (miRNAs) and endoplasmic reticulum (ER) stress play critical roles in myoblast differentiation. However, the regulation roles of miRNAs and ER stress in myogenic differentiation have not been fully revealed and need to be further studied. Here, we discovered that the expression levels of miR-181a-5p were strongly upregulated during C2C12 cell differentiation. miR-181a-5p overexpression promoted ER stress and differentiation of C2C12 cells, which was accompanied by increasing expression levels of marker genes related to ER stress-mediated apoptosis and myogenic differentiation. Opposite results were observed after inhibition of the miR-181a-5p expression. The gain- and loss-of-function experiments on C2C12 cells showed that miR-181a-5p affected the development of muscle fiber type, but had no significant influence on C2C12 cell proliferation. In the ER-stressed C2C12 cells induced by thapsigargin (Tg), the expression levels of both miR-181a-5p and marker genes related to ER stress and myogenesis were upregulated. In the ER-stressed C2C12 cells and porcine muscle fibroblast (PMF) cells pretreated with Tg, we found that miR-181a-5p targeted glucose-regulated protein, 78 kDa/binding immunoglobulin protein (GRP78/BIP), and influenced cell apoptosis. In conclusion, these results indicate that miR-181a-5p and ER stress have positive synergistic effects on myogenic differentiation by increasing the expression levels of myogenic differentiation key genes and activating the ER stress-mediated apoptosis signaling pathway. [ABSTRACT FROM AUTHOR]

The mutant p53 plays a vital role in the control of cell survival and division under various stresses, including apoptosis and ferroptosis. Here, we showed that eupaformosanin (Eup), a natural compound isolated from Eupatorium cannabinum Linn., significantly inhibited the viability of triple-negative breast cancer (TNBC) cells. Meanwhile, mitochondrial apoptosis contributed to the apoptosis induced by Eup, followed by the disruption of mitochondrial membrane potential (MMP; Δψm) and accumulation of mitochondrial ROS (mt ROS). Apoptosis inhibitor Z-VAD rescued Eup-induced cell death. Afterward, ferroptosis-induced cell death was demonstrated after treatment with Eup, accompanied by lipid reactive oxygen species (ROS) accumulation, glutathione (GSH) depletion, and iron increase. These events were blocked by ferroptosis inhibitors ferrostatin-1 (Fer-1), deferoxamine (DFO), and liproxstatin-1 (lip-1), indicating that ferroptosis facilitated Eup-induced cell death. Furthermore, Eup regulated mutant p53 ubiquitination. Mutant p53 signaling pathway participated in Eup-induced apoptosis and ferroptosis, which were rescued when mutant p53 was silent in TNBC cells. Also, Eup exerted an anti-TNBC effect by inducing apoptosis and ferroptosis in vivo. Taken together, the data demonstrate that the natural compound Eup is a potential TNBC therapeutic agent that induces apoptosis and ferroptosis through ubiquitination of mutant p53. [Display omitted] [ABSTRACT FROM AUTHOR]

The alleviation of the impact of sucrose treatment on programmed cell death (PCD) in postharvest broccoli has been reported in previous studies; however, the fundamental mechanisms are still not fully understood. This study found that exogenous sucrose treatment maintained the mitochondrial structure in broccoli buds, enhanced mitochondrial membrane potential (ΔΨm), elevated activity of antioxidant enzymes and decreased levels of reactive oxygen species (ROS). These changes ultimately inhibited the release of Cytochrome c/a (Cytc/a) and delayed PCD in broccoli buds. Furthermore, the expression of BoVDAC2 progressively decreased with prolonged storage. To further explore the relationship between BoVDAC2 and PCD, BoVDAC2 was overexpressed in BY-2 cells in this study. Compared to wild-type cells, BoVDAC2 -overexpressing cells showed reduced mitochondrial ultrastructure damage, increased cell vitality, enhanced antioxidant enzyme activity, and decreased ROS production. Additionally, BoVDAC2 overexpression elevated energy metabolism, maintained mitochondrial membrane integrity, inhibited Cytc/a release, suppressed caspase-like activity, and improved the expression of genes related to PCD in BY-2 cells. Overall, the findings suggest that sucrose induces BoVDAC2 expression, which helps to maintain mitochondrial structure and function in postharvest broccoli buds, thereby mitigating PCD. [Display omitted] • Sucrose treatment delayed the yellowing process of broccoli buds. • Increase the expression of BoVDAC2 in broccoli with sucrose treatment. • BoVDAC2 in BY-2 cells reduced the level of oxidative stress. • BoVDAC2 in BY-2 cells delayed the occurrence of PCD. [ABSTRACT FROM AUTHOR]

The beneficial effects of γ-aminobutyric acid (GABA) on oxidative damage and programmed cell death (PCD) in fresh-cut pumpkins were preliminarily demonstrated in our previous research. However, the mechanism by which GABA alleviates PCD in fresh-cut pumpkins remains unclear. In this study, we discovered that GABA treatment effectively reduced mitochondrial structural damage and membrane permeability while increasing the mitochondrial membrane potential (ΔΨm) in fresh-cut pumpkins compared to the control. Additionally, GABA-treated pumpkins exhibited higher adenosine triphosphate (ATP) content and energy charge, consistent with increased activity of H+-ATPase, succinate dehydrogenase (SDH), malate dehydrogenase (MDH), and cytochrome c oxidase (COX). Conversely, the activity of alternative oxidase (AOX), Ca2+-ATPase, and K+-ATPase was reduced in GABA-treated pumpkins compared to the control. Moreover, key factors in apoptosis, such as the activity of six caspases and the gene expression of apoptosis-inducing factors (CmAIFs) and metacaspases (CmMCAs), were significantly decreased following GABA treatment. In conclusion, GABA effectively suppressed PCD in postharvest fresh-cut pumpkins by improving mitochondrial physiological properties and energy metabolism while inhibiting apoptosis-related factors. [Display omitted] ● GABA maintained normal ΔΨm, MPTP and ultrastructure of mitochondria. ● GABA increased energy charge by enhancing TCA cycle and electron transport chain. ● GABA decreased the activity and gene expression of apoptosis factors to inhibit PCD. [ABSTRACT FROM AUTHOR]

Programmed cell death (PCD) is a genetically regulated process of cell suicide essential for plant development. The 'malate valve' is a mechanism that ensures redox balance across different subcellular compartments. In broccoli, the BomMDH1 gene encodes malate dehydrogenase in mitochondria, a critical enzyme in the 'malate circulation' pathway. This study investigates the functional role of BomMDH1 in malate (MA)-induced apoptosis in bright yellow-2 (BY-2) suspension cells. Findings revealed that transgenic cells overexpressing BomMDH1 showed enhanced viability under MA-induced oxidative stress compared to wild-type (WT) cells. Overexpression of BomMDH1 also reduced levels of reactive oxygen species (ROS), hydrogen peroxide (H 2 O 2), and malondialdehyde (MDA), while increasing the expression of antioxidant enzyme genes such as NtAPX , NtAOX1a , NtSOD , and NtMDHAR. Additionally, treatment with salicylhydroxamic acid (SHAM), a characteristic inhibitor of mitochondrial respiration, further improved the anti-apoptotic activity of BY-2 cells. Overall, these results highlighted the function of the BomMDH1 gene and the potential of SHAM treatment in mitigating oxidative stress in BY-2 suspension cells. • Malate treatment accelerated the yellowing process of broccoli. • Overexpressing BomMDH1 in BY-2 cells exhibited stronger cell viability. • Confirmed the effective role of SHAM in BY-2 suspension cells exposed to MA. [ABSTRACT FROM AUTHOR]

The regulation of γ-aminobutyric acid (GABA) on plant anti-oxygenation and programmed cell death (PCD) in fresh-cut pumpkins was investigated. Exogenous GABA positively promoted GABA accumulation and alleviated oxidant damage in pumpkins tissue. Pumpkins treated with GABA showed lower electrolyte leakage, reactive oxygen species (ROS) and MDA content, while higher activities of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), and ascorbate peroxidase (APX) as compared to the non-treated tissues. Our results also found that GABA treatment retarded DNA degradation and cytochrome c release and delayed the apoptosis of pumpkin cells. On the other hand, the inhibitor of GABA generation, 3-mercaptopropionic acid (3-MP) treatment not only accelerated oxidant damage, but also induced cell death involving chromatin condensation, DNA ladder and cytochrome c releasing. Taken together, our present research indicated that exogenous GABA could alleviate the wound-induced oxidative stress and PCD occurrence in fresh-cut pumpkins. [Display omitted] • Exogenous GABA alleviated oxidant damage in fresh-cut pumpkins. • 3-Mercaptopropionic acid treatment accelerated oxidant damage and induced PCD. • GABA involved the PCD event and delayed PCD process in fresh-cut pumpkins. [ABSTRACT FROM AUTHOR]

Plant senescence is the final period of the plant life cycle and represents a programmed cell death (PCD) process. Melatonin, a versatile signaling molecule, exhibits potent free radical scavenging properties and actively participates in cellular antioxidant systems, effectively retarding plant senescence throughout its development. In order to illustrate the impact of melatonin on PCD during senescence, we examined the effects of exogenous melatonin at 80 μmol L−1 on PCD and mitochondrial function in broccoli bud cells at 25 ℃. The findings revealed that melatonin treatment not only maintained the morphological integrity of chloroplasts and mitochondria but also attenuated the decline in chlorophyll content by reducing the expression of genes that metabolize chlorophyll, including BoPPH , BoPAO , and BoSGR1. In comparison to the control group, the treatment decreased the content of H 2 O 2 and the rate of O 2 - production in mitochondria and simultaneously enhanced the activities of ascorbate peroxidase (APX), catalase (CAT), and superoxide dismutase (SOD), resulting in effective maintenance of reactive oxygen species (ROS) homeostasis and reduction of the level of lipid peroxidation in broccoli buds during senescence. Meanwhile, exogenous melatonin treatment up-regulated alternative oxidase (AOX) activity and its gene expression while delayed the reduction of cytochrome oxidase (COX) activity and restrained the over-opening of mitochondrial permeability transition pores (MPTP). Additionally, exogenous melatonin decelerated the increase in membrane potential and prevented mitochondria from releasing excess cytochrome c (Cyt c) into the cytoplasm. Taken together, the results indicate that melatonin treatment delayed mitochondrial dysfunction, preserved mitochondrial ROS homeostasis, and consequently postponed the initiation of PCD, ultimately inhibiting senescence in broccoli. ● Exogenous melatonin protects the structural of chloroplasts and mitochondrial. ● Exogenous melatonin down-regulates the chlorophyll degrading gene. ● Exogenous melatonin reduces mitochondrial dysfunction in broccoli. ● Exogenous melatonin delays the decline of APX, CAT and SOD activities. ● Exogenous melatonin delays the occurrence of programmed cell death in broccoli. [ABSTRACT FROM AUTHOR]

It is well established that programmed cell death (PCD) occurred in broccoli during postharvest senescence, but no studies have been conducted on the regulation of broccoli cytochrome f by mannose treatment and its relationship with PCD. In this study, we treated broccoli buds with mannose to investigate the changes in color, total chlorophyll content, gene expression related to chlorophyll metabolism, chloroplast structure, and cytochrome f determination during postharvest storage. In addition, to investigate the effect of cytochrome f on PCD, we extracted cytochrome f from broccoli and treated Nicotiana tabacum L. cv Bright Yellow 2 (BY-2) cells with extracted cytochrome f from broccoli at various concentrations. The results showed that cytochrome f can induce PCD in tobacco BY-2 cells, as evidenced by altered cell morphology, nuclear chromatin disintegration, DNA degradation, decreased cell viability, and increased caspase-3-like protease production. Taken together, our study indicated that mannose could effectively delay senescence of postharvest broccoli by inhibiting the expression of gene encoding cytochrome f which could induce PCD. • Mannose treatment delayed the senescence process of broccoli. • Mannose maintained chloroplast structure and mitigated degradation in broccoli. • Cytochrome f from broccoli induced PCD in BY-2 cells. [ABSTRACT FROM AUTHOR]

• Mannose delayed the decline of chlorophyll content. • Mannose regulated the gene expressions of chlorophyll catabolic enzymes. • Mannose retarded the PCD in broccoli by a mitochondria-dependent pathway. The effect of mannose treatment on chlorophyll catabolism and programmed cell death (PCD) in broccoli florets during storage at 20 ℃ has been investigated. The results showed that mannose treatment efficiently restrained chlorophyll degradation in broccoli florets via regulating chlorophyll catabolism genes (BoPPH, BoPAO, BoRCCR, BoSGR1 and BoSGR2). Meanwhile, mannose treatment efficiently suppressed the occurrence of PCD through the mitochondria-dependent pathway in broccoli florets. Less reactive oxygen species content observed in mannose-treated broccoli florets delayed the increase of mitochondrial membrane potential and inhibited unfolding of mitochondria permeability transition pore, which was associated with the inhibition of cytochrome c liberation in mitochondria and the maintenance of high levels of cytochrome c/a and the ratio of Ca2+ content, the activities of cytochrome c oxidase and alternative oxidase. These results suggested that mannose treatment inhibited senescence of broccoli florets through suppressing the chlorophyll degradation and the appearance of PCD. [ABSTRACT FROM AUTHOR]

• Broccoli senescence occurred with PCD and sucrose delayed the PCD process. • Sucrose inhibited DNA degradation and delayed the early apoptosis of broccoli cell; • Caspase-like activities in broccoli cells were suppressed under sucrose treatment. The effect of sucrose treatment on morphological and biochemical features of programmed cell death (PCD) in broccoli florets was explored. The apoptosis of untreated and treated cells was evaluated by flow cytometry (FCM) and by laser scanning confocal microscopy (LSCM). DNA ladders, resulting from the cleavage of nuclear DNA into oligonucleosomal fragments in PCD, were evaluated by gel electrophoresis. DNA frag-mentation and nuclear DNA condensation were further confirmed by using the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end in situ labeling (TUNEL). The results revealed that sucrose treatment inhibited DNA degradation, and delayed the early apoptosis of broccoli cell as demonstrated with FCM and LSCM. DNA laddering was observed in control broccoli florets, which could not be detected in sucrose-treated samples. In addition, the caspase-like activities in broccoli cells were suppressed under sucrose treatment. Taken together, our results suggested that PCD was delayed in sucrose-treated broccoli florets, which provided a new model to investigate the mechanism of sugar in regulating PCD during postharvest storage. [ABSTRACT FROM AUTHOR]

• Sucrose treatment retarded the onset of PCD in broccoli florets. • Sucrose significantly retarded the cytochrome c release from mitochondria. • Sucrose improved the mitochondrial physiological properties. The effect of sucrose treatment on mitochondrial membrane potential (ΔΨm), contents of reactive oxygen species (ROS), cytochrome c oxidase (COX) activity, alternative oxidase (AOX) activity, mitochondrial Ca2+ content and cytochrome c release from mitochondria in broccoli florets during storage at 20 °C was investigated. The results showed that sucrose treatment effectively retarded the onset of programmed cell death (PCD) in broccoli florets by a mitochondria-dependent pathway. Sucrose treatment inhibited the increase of ROS and a decrease in ΔΨm, which was associated with the open state of permeability transition pore (PTP) and the ratio of cytochrome c/a. The decrease of COX activity and Ca2+ content in broccoli florets was suppressed under sucrose treatment. Sucrose treatment maintained higher AOX activity, and retarded the cytochrome c release from mitochondria. These results indicated that sucrose treatment delayed the appearance of PCD in broccoli florets through improving mitochondrial physiological properties. [ABSTRACT FROM AUTHOR]