Accelerated Cell Death Research Articles

A single fluorescent probe to examine the dynamics of mitochondria-lysosome interplay and extracellular vesicle role in ferroptosis

Ferroptosis is a non-apoptotic form of cell death characterized by iron-dependent lipid peroxidation and glutathione (GSH) depletion. Despite recent advances, challenges remain in understanding the bidirectional interactions or interplay between organelles during ferroptosis. In this study, we aimed to understand the interplay between mitochondria (Mito) and lysosomes (Lyso) in cell homeostasis and ferroptosis. For this purpose, we designed a single fluorescent probe that marks GSH in Mito and hypochlorous acid (HOCl) in Lyso with two distinct emissions. Using this dual-targeted single fluorescent probe (9-morphorino pyronine), we detected Mito-Lyso interplay in ferroptosis. We disclosed differences in Mito-Lyso interplay depending on the induction of ferroptosis. Although erastin treatment decreased GSH, RSL3 triggered a HOCl burst, andFIN56- and FINO2-induced ferroptosis increased GSH and HOCl. Additionally, we showed that only extracellular vesicles generated during erastin-induced ferroptosis could spontaneously move and dock to neighboring cells, resulting in accelerated cell death.

Potential Role of Endoplasmic Reticulum Stress in Modulating Protein Homeostasis in Oligodendrocytes to Improve White Matter Injury in Preterm Infants.

Preterm white matter injury (WMI) is a demyelinating disease with high incidence and mortality in premature infants. Oligodendrocyte cells (OLs) are a specialized glial cell that produces myelin proteins and adheres to the axons providing energy and metabolic support which susceptible to endoplasmic reticulum protein quality control. Disruption of cellular protein homeostasis led to OLs dysfunction and cell death, immediately, the unfolded protein response (UPR) activated to attempt to restore the protein homeostasis via IRE1/XBP1s, PERK/eIF2α and ATF6 pathway that reduced protein translation, strengthen protein-folding capacity, and degraded unfolding/misfolded protein. Moreover, recent works have revealed the conspicuousness function of ER signaling pathways in regulating influenced factors such as calcium homeostasis, mitochondrial reactive oxygen generation, and autophagy activation to regulate protein hemostasis and improve the myelination function of OLs. Each of the regulation modes and their corresponding molecular mechanisms provides unique opportunities and distinct perspectives to obtain a deep understanding of different actions of ER stress in maintaining OLs' health and function. Therefore, our review focuses on summarizing the current understanding of ER stress on OLs' protein homeostasis micro-environment in myelination during white matter development, as well as the pathophysiology of WMI, and discussing the further potential experimental therapeutics targeting these factors that restore the function of the UPR in OLs myelination function.

SIMULATION OF PHYSIOLOGICAL AND DETRIMENTAL LOADING IN WHOLE INTERVERTEBRAL DISC ORGAN MODELS

Mechanical loading is important to maintain the homeostasis of the intervertebral disc (IVD) under physiological conditions but can also accelerate cell death and tissue breakdown in a degenerative state. Bioreactor loaded whole organ cultures are instrumental for investigating the effects of the mechanical environment on the IVD integrity and for preclinical testing of new therapies under simulated physiological conditions. Thereby the loading parameters that determine the beneficial or detrimental reactions largely depend on the IVD model and its preparation. Within this symposium we are discussing the use of bovine caudal IVD culture models to reproduce tissue inflammation or matrix degradation with or without bioreactor controlled mechanical loading. Furthermore, the outcome parameters that define the degenerative state of the whole IVD model will be outlined. Besides the disc height, matrix integrity, cell viability and phenotype expression, the tissue secretome can provide indications about potential interactions of the IVD with other cell types such as neurons. Finally, a novel multiaxial bioreactor setup capable of mimicking the six degrees-of-freedom loading environment of IVDs will be introduced that further advances the relevance of preclinical ex-vivo testing.

Berberine mediated fluorescent gold nanoclusters in biomimetic erythrocyte ghosts as a nanocarrier for enhanced photodynamic treatment.

Photodynamic therapy (PDT) is a well-established cancer treatment method that employs light to generate reactive oxygen species (ROS) causing oxidative damage to cancer cells. Nevertheless, PDT encounters challenges due to its oxygen-dependent nature, which makes it less effective in hypoxic tumor environments. To address this issue, we have developed a novel nanocomposite known as AuNC@BBR@Ghost. This nanocomposite combines the advantageous features of erythrocyte ghost membranes, the photoresponsive properties of gold nanoclusters (AuNC) and the anticancer characteristics of Berberine (BBR) for cancer treatment. Our synthesized AuNC efficiently produce ROS, with a 25% increase in efficiency when exposed to near-infrared (NIR) irradiation. By harnessing the oxygen-carrying capacity of erythrocyte ghost cells, AuNC@BBR@Ghost demonstrates a significant improvement in ROS generation, achieving an 80% efficiency. Furthermore, the AuNC exhibit tunable emission wavelengths due to their excellent fluorescent properties. In normoxic conditions, treatment of A549 lung carcinoma cells with AuNC@BBR@Ghost followed by exposure to 808 nm NIR irradiation results in a notable increase in intracellular ROS levels, accelerating cell death. In hypoxic conditions, when A549 cells were treated with AuNC@BBR@Ghost, the erythrocyte ghost acted as an oxygen supplement due to the residual hemoglobin, alleviating hypoxia and enhancing the nanocomposite's sensitivity to PDT treatment. Thus, the AuNC@BBR@Ghost nanocomposite achieves an improved effect by combining the advantageous properties of its individual components, resulting in enhanced ROS generation and adaptability to hypoxic conditions. This innovative approach successfully overcomes PDT's limitations, making AuNC@BBR@Ghost a promising nanotheranostic agent with significant potential for advanced cancer therapy.

(Nano)biotechnological approaches in the treatment of cervical cancer: integration of engineering and biology.

Cervical cancer is one of the most malignant gynaecological tumors characterised with the aggressive behaviour of the tumor cells. In spite of the development of different strategies for the treatment of cervical cancer, the tumor cells have developed resistance to conventional therapeutics. On the other hand, nanoparticles have been recently applied for the treatment of human cancers through delivery of drugs and facilitate tumor suppression. The stimuli-sensitive nanostructures can improve the release of therapeutics at the tumor site. In the present review, the nanostructures for the treatment of cervical cancer are discussed. Nanostructures can deliver both chemotherapy drugs and natural compounds to increase anti-cancer activity and prevent drug resistance in cervical tumor. Moreover, the genetic tools such as siRNA can be delivered by nanoparticles to enhance their accumulation at tumor site. In order to enhance selectivity, the stimuli-responsive nanoparticles such as pH- and redox-responsive nanocarriers have been developed to suppress cervical tumor. Moreover, nanoparticles can induce photo-thermal and photodynamic therapy to accelerate cell death in cervical tumor. In addition, nanobiotechnology demonstrates tremendous potential in the treatment of cervical cancer, especially in the context of tumor immunotherapy. Overall, metal-, carbon-, lipid- and polymer-based nanostructures have been utilized in cervical cancer therapy. Finally, hydrogels have been developed as novel kinds of carriers to encapsulate therapeutics and improve anti-cancer activity.

Reduction of HIF-1α/PD-L1 by Catalytic Topoisomerase Inhibitor Induces Cell Death Through Caspase Activation in Cancer Cells Under Hypoxia.

Under severe hypoxia, cellular apoptosis is induced through hypoxia-inducible factor 1, alpha subunit (HIF-1α)-dependent P53 accumulation and P53 phosphorylation via ataxia telangiectasia mutated and ataxia telangiectasia and RAD3-related (ATR) activation via replication stress-induced DNA damage response (DDR) activation. We previously demonstrated that the topoisomerase I catalytic inhibitor, 3-O-(2'E,4'Z-decadienoyl)-20-O-acetylingenol (3EZ,20Ac-ingenol), induced apoptosis in Jeko-1 and Panc-1 cells, both of which show cyclin D1 overexpression. After progression to the S phase facilitated by nuclear cyclin D1, an intra S phase checkpoint was induced in the presence of 3EZ,20Ac-ingenol, by ATR activation in response to replication stress-induced DDR. In this study, we examined whether 3-O-(2'E,4'E-decadienoyl)-20-O-acetylingenol (3EE,20Ac-ingenol) might induce a higher degree of P53 phosphorylation and additional HIF-1α and P53 accumulation in response to replication stress-induced DDR activation under hypoxic conditions than under normoxic conditions, by controlling ATR activation. In the Panc-1 cells, 3EE,20Ac-ingenol induced P53 activation and HIF-1α-dependent P53 accumulation through cooperative ATR activation via hypoxia-induced DDR activation. Jeko-1 cells showed slight HIF-1α accumulation under hypoxia, but HIF-1α-dependent 53 accumulation was not observed in the presence of 3EE,20Ac-ingenol, so that the cells remained resistant to hypoxia. 3EE,20Ac-ingenol induces an intricate interplay between P53 and HIF-1α accumulation via ATR activation that results in a high P53 accumulation, which promoted transient expression and early disappearance of HIF-1α, accelerating cell death. Strong P53 accumulation and consequent phosphatase and tensin homolog deleted on chromosome 10 activation in Panc-1 cells also reduced HIF-1α accumulation and programmed death-ligand 1 expression, which resulted in intense apoptosis.

Aberrant accumulation of ceramides in mitochondria triggers cell death by inducing autophagy in Arabidopsis.

Sphingolipids are membrane lipids and play critical roles in signal transduction. Ceramides are central components of sphingolipid metabolism that are involved in cell death. However, the mechanism of ceramides regulating cell death in plants remains unclear. Here, we found that ceramides accumulated in mitochondria of accelerated cell death 5 mutant (acd5), and expression of mitochondrion-localized ceramide kinase (ACD5) suppressed mitochondrial ceramide accumulation and the acd5 cell death phenotype. Using immuno-electron microscopy, we observed hyperaccumulation of ceramides in acer acd5 double mutants, which are characterized by mutations in both ACER (alkaline ceramidase) and ACD5 genes. The results confirmed that plants with specific ceramide accumulation exhibited localization of ceramides to mitochondria, resulting in an increase in mitochondrial reactive oxygen species production. Interestingly, when compared with the wild type, autophagy-deficient mutants showed stronger resistance to ceramide-induced cell death. Lipid profiling analysis demonstrated that plants with ceramide accumulation exhibited a significant increase in phosphatidylethanolamine levels. Furthermore, exogenous ceramide treatment or endogenous ceramide accumulation induces autophagy. When exposed to exogenous ceramides, an increase in the level of the autophagy-specific ubiquitin-like protein, ATG8e, associated with mitochondria, where it directly bound to ceramides. Taken together, we propose that the accumulation of ceramides in mitochondria can induce cell death by regulating autophagy.

Unveiling autophagy complexity in leukemia: The molecular landscape and possible interactions with apoptosis and ferroptosis

Autophagy is a self-digestion multistep process in which causes the homeostasis through degradation of macromolecules and damaged organelles. The autophagy-mediated tumor progression regulation has been a critical point in recent years, revealing the function of this process in reduction or acceleration of carcinogenesis. Leukemia is a haematological malignancy in which abnormal expansion of hematopoietic cells occurs. The current and conventional therapies from chemotherapy to cell transplantation have failed to appropriately treat the leukemia patients. Among the mechanisms dysregulated in leukemia, autophagy is a prominent one in which can regulate the hallmarks of this tumor. The protective autophagy inhibits apoptosis and ferroptosis in leukemia, while toxic autophagy accelerates cell death. The proliferation and invasion of tumor cells are tightly regulated by the autophagy. The direction of regulation depends on the function of autophagy that is protective or lethal. The protective autophagy accelerates chemoresistance and radio-resistsance. The non-coding RNAs, histone transferases and other pathways such as PI3K/Akt/mTOR are among the regulators of autophagy in leukemia progression. The pharmacological intervention for the inhibition or induction of autophagy by the compounds including sesamine, tanshinone IIA and other synthetic compounds can chance progression of leukemia.

APOL1-G2 accelerates nephrocyte cell death by inhibiting the autophagy pathway.

People of African ancestry who carry the APOL1 risk alleles G1 or G2 are at high risk of developing kidney diseases through not fully understood mechanisms that impair the function of podocytes. It is also not clear whether the APOL1-G1 and APOL1-G2 risk alleles affect these cells through similar mechanisms. Previously, we have developed transgenic Drosophila melanogaster lines expressing either the human APOL1 reference allele (G0) or APOL1-G1 specifically in nephrocytes, the cells homologous to mammalian podocytes. We have found that nephrocytes that expressed the APOL1-G1 risk allele display accelerated cell death, in a manner similar to that of cultured human podocytes and APOL1 transgenic mouse models. Here, to compare how the APOL1-G1 and APOL1-G2 risk alleles affect the structure and function of nephrocytes in vivo, we generated nephrocyte-specific transgenic flies that either expressed the APOL1-G2 or both G1 and G2 (G1G2) risk alleles on the same allele. We found that APOL1-G2- and APOL1-G1G2-expressing nephrocytes developed more severe changes in autophagic pathways, acidification of organelles and the structure of the slit diaphragm, compared to G1-expressing nephrocytes, leading to their premature death. We conclude that both risk alleles affect similar key cell trafficking pathways, leading to reduced autophagy and suggesting new therapeutic targets to prevent APOL1 kidney diseases.

Small proteins modulate ion-channel-like ACD6 to regulate immunity in Arabidopsis thaliana

The multi-pass transmembrane protein ACCELERATED CELL DEATH 6 (ACD6) is an immune regulator in Arabidopsis thaliana with an unclear biochemical mode of action. We have identified two loci, MODULATOR OF HYPERACTIVE ACD6 1 (MHA1) and its paralog MHA1-LIKE (MHA1L), that code for ∼7kDa proteins, which differentially interact with specific ACD6 variants. MHA1L enhances the accumulation of an ACD6 complex, thereby increasing the activity of the ACD6 standard allele for regulating plant growth and defenses. The intracellular ankyrin repeats of ACD6 are structurally similar to those found in mammalian ion channels. Several lines of evidence link increased ACD6 activity to enhanced calcium influx, with MHA1L as a direct regulator of ACD6, indicating that peptide-regulated ion channels are not restricted to animals.

Developmental and foliation changes due to dysregulation of adenosine kinase in the cerebellum

Adenosine kinase (ADK), the major adenosine-metabolizing enzyme, plays a key role in brain development and disease. In humans, mutations in the Adk gene have been linked to developmental delay, stunted growth, and intellectual disability. To better understand the role of ADK in brain development, it is important to dissect the specific roles of the two isoforms of the enzyme expressed in the cytoplasm (ADK-S) and cell nucleus (ADK-L). We, therefore, studied brain development in Adk-tg transgenic mice, which only express ADK-S in the absence of ADK-L throughout development. In the mutant animals, we found a reduction in the overall brain, body size, and weight during fetal and postnatal development. As a major developmental abnormality, we found a profound change in the foliation pattern of the cerebellum. Strikingly, our results indicated aberrant Purkinje cells arborization at P9 and accelerated cell death at P6 and P9. We found defects in cerebellar cell proliferation and migration using a bromodeoxyuridine (BrdU)-based cell proliferation assay at postnatal day 7. Our data demonstrate that dysregulation of ADK expression during brain development profoundly affects brain growth and differentiation.

FRIENDLY is required for efficient dark-induced mitophagy and controlled senescence in Arabidopsis.

Mitochondria play essential roles in plant metabolism, supporting both development and stress responses. To maintain a healthy pool of mitochondria, several quality control systems are in place. Selected degradation of mitochondria by autophagy -mitophagy- has been extensively studied in yeast and animals, but information on mitophagy components in plants is limited. The 'Friendly Mitochondria' (FRIENDLY; FMT) protein, homologous to 'clustered mitochondria protein homolog' CLU in animals, was recently suggested to mediate mitophagy of depolarized mitochondria. In this study, we evaluated the role of FMT in carbon starvation- and dark senescence-induced mitophagy in Arabidopsis. Using mitophagy flux assays, we show that loss of FMT results in decreased mitophagy during dark-induced senescence. Mitophagy induced by inhibition of Target of Rapamycin (TOR) signalling is also partially impaired in fmt mutants. The FMT protein is mostly localised in the cytosol, but we show that during darkness FMT is redistributed into speckles, some of which associate with mitochondria. Fmt mutants were initially identified for their abnormal mitochondrial morphology, with mitochondria often forming clusters. We found that dark senescence strongly increases the number and size of mitochondrial clusters in fmt mutants. In agreement with a role for FMT in mitophagy, we show that fmt mutants show accelerated senescence phenotypes comparable to autophagy 11 (atg11) mutants, but less prominent than in atg5 mutants. Furthermore, FMT prevents excessive dark-induced cell death and hydrogen peroxide production, and supports mitophagy and greening in etiolated seedlings. Our findings thus indicate that FMT contributes to mitophagy and provide evidence that mitophagy is required for controlled senescence and prevention of accelerated cell death. We propose that FMT mediates efficient mitophagy by preventing mitochondrial clustering, thereby allowing mitochondria to be captured more effectively by autophagosomes, rather than by acting as a direct mitophagy receptor.

During heat stress in Myxococcus xanthus, the CdbS PilZ domain protein, in concert with two PilZ-DnaK chaperones, perturbs chromosome organization and accelerates cell death.

C-di-GMP is a bacterial second messenger that regulates diverse processes in response to environmental or cellular cues. The nucleoid-associated protein (NAP) CdbA in Myxococcus xanthus binds c-di-GMP and DNA in a mutually exclusive manner in vitro. CdbA is essential for viability, and CdbA depletion causes defects in chromosome organization, leading to a block in cell division and, ultimately, cell death. Most NAPs are not essential; therefore, to explore the paradoxical cdbA essentiality, we isolated suppressor mutations that restored cell viability without CdbA. Most mutations mapped to cdbS, which encodes a stand-alone c-di-GMP binding PilZ domain protein, and caused loss-of-function of cdbS. Cells lacking CdbA and CdbS or only CdbS were fully viable and had no defects in chromosome organization. CdbA depletion caused post-transcriptional upregulation of CdbS accumulation, and this CdbS over-accumulation was sufficient to disrupt chromosome organization and cause cell death. CdbA depletion also caused increased accumulation of CsdK1 and CsdK2, two unusual PilZ-DnaK chaperones. During CdbA depletion, CsdK1 and CsdK2, in turn, enabled the increased accumulation and toxicity of CdbS, likely by stabilizing CdbS. Moreover, we demonstrate that heat stress, possibly involving an increased cellular c-di-GMP concentration, induced the CdbA/CsdK1/CsdK2/CdbS system, causing a CsdK1- and CsdK2-dependent increase in CdbS accumulation. Thereby this system accelerates heat stress-induced chromosome mis-organization and cell death. Collectively, this work describes a unique system that contributes to regulated cell death in M. xanthus and suggests a link between c-di-GMP signaling and regulated cell death in bacteria.

‘Candidatus Liberibacter asiaticus’ secretory protein SDE3 inhibits host autophagy to promote Huanglongbing disease in citrus

ABSTRACT Antimicrobial acroautophagy/autophagy plays a vital role in degrading intracellular pathogens or microbial molecules in host-microbe interactions. However, microbes evolved various mechanisms to hijack or modulate autophagy to escape elimination. Vector-transmitted phloem-limited bacteria, Candidatus Liberibacter (Ca. Liberibacter) species, cause Huanglongbing (HLB), one of the most catastrophic citrus diseases worldwide, yet contributions of autophagy to HLB disease proliferation remain poorly defined. Here, we report the identification of a virulence effector in “Ca. Liberibacter asiaticus” (Las), SDE3, which is highly conserved among the “Ca. Liberibacter”. SDE3 expression not only promotes the disease development of HLB and canker in sweet orange (Citrus sinensis) plants but also facilitates Phytophthora and viral infections in Arabidopsis, and Nicotiana benthamiana (N. benthamiana). SDE3 directly associates with citrus cytosolic glyceraldehyde-3-phosphate dehydrogenases (CsGAPCs), which negatively regulates plant immunity. Overexpression of CsGAPCs and SDE3 significantly inhibits autophagy in citrus, Arabidopsis, and N. benthamiana. Intriguingly, SDE3 undermines autophagy-mediated immunity by the specific degradation of CsATG8 family proteins in a CsGAPC1-dependent manner. CsATG8 degradation is largely rescued by treatment with an inhibitor of the late autophagic pathway, E64d. Furthermore, ectopic expression of CsATG8s enhances Phytophthora resistance. Collectively, these results suggest that SDE3-CsGAPC interactions modulate CsATG8-mediated autophagy to enhance Las progression in citrus. Abbreviations: ACP: asian citrus psyllid; ACD2: ACCELERATED CELL DEATH 2; ATG: autophagy related; Ca. Liberibacter: Candidatus Liberibacter; CaMV: cauliflower mosaic virus; CMV: cucumber mosaic virus; Cs: Citrus sinensis; EV: empty vector; GAPC: cytosolic glyceraldehyde-3-phosphate dehydrogenase; HLB: huanglongbing; H2O2: hydrogen peroxide; Las: liberibacter asiaticus; Laf: liberibacter africanus; Lam: liberibacter americanus; Pst: Pseudomonas syringae pv. tomato; PVX: potato virus X; ROS: reactive oxygen species; SDE3: sec-delivered effector 3; TEM: transmission electron microscopy; VIVE : virus-induced virulence effector; WT: wild-type; Xcc: Xanthomonas citri subsp. citri;

Autophagy accelerates cell death after desiccation and hydration stress in Physcomitrium

Autophagy is the process responsible for degrading cytoplasmic components in lysosomes and vacuoles. Autophagy-deficient mutants are generally more sensitive to environmental stress than their wild-type (WT) plants. In this study, however, we found that autophagy-deficient (atg) mutants of Physcomitrium, in which an autophagy-related gene, either ATG5 or ATG7, is disrupted, are more tolerant to extreme desiccation stress than WT plants. The colonies were treated with the phytohormone abscisic acid, desiccated using silica gel, and hydrated on a nutrient-sufficient medium. atg mutant colonies survived this stress program and produced green colonies whereas the WT colonies did not. Evans Blue staining revealed obvious cell death in both colonies, but the rate was higher in WT than in atg5 colonies. atg cells had a higher protein content, accumulated more sucrose, and had greater water-holding capacity than WT cells. These factors likely reduce the rate of cell death with less damage during the stress program. On the other hand, inhibition of autophagy during the hydration step with the inhibitor 3-methyladenine increased survival, suggesting that autophagy occurring during the hydration step is involved in cell death. We conclude that Physcomitrium autophagy-deficient cells are more tolerant to extreme desiccation stress than WT cells due to higher water-holding capacity and the promotion of cell death by autophagy through an unknown mechanism. These findings challenge the conventional understanding of the role of autophagy in cell survival against stress and highlight the importance of further research in this area.

A Curcumin‐Based MOF Embedded Ag NPs and Modified with Polydopamine for Dual‐Drug Delivery and Dual Biological Window Responsive Synergetic Cancer Therapy

AbstractA dual‐drug delivery, pH‐responsive composite nanoplatform (MAPD NPs) that can respond to two biological windows is developed to improve the efficacy of synergetic chemotherapic/photothermal/chemodynamic therapy (CDT) against tumors. This nanoplatform is surface‐modified polydopamine (PDA) with excellent biocompatibility as the shell and Ag NPs as the catalyst for CDT. The curcumin (Cur) acts as an organic ligand to be encapsulated in metal−biomolecule frameworks (Bio‐MOFs) by self‐assembly, and Bio‐MOF acts as a delivery carrier to deliver of DOX•HCl and then releases the Cur when it degrades in vivo. Moreover, Bio‐MOF can be taken up by cells faster and accelerate cell death compared to free Cur. PDA modification enables MAP (PDA@MOF‐Ag) to have photothermal properties under 808 and 1064 nm light irradiation, which not only improves the biocompatibility of MAP but also makes it produce high heat and abundant ·OH. The photothermal performance of MAP is stable after irradiation at 808 or 1064 nm, and the photothermal conversion efficiency reaches 63.57% and 26.25%. The survival rate of HeLa cells co‐incubation with MAPD NPs after irradiation at 808 and 1064 nm decreases to 19.52 ± 0.69% and 30.48 ± 0.49%, respectively, providing a feasible scheme for the realization of deep tumor killing.

Extracellular ATP accelerates cell death and decreases tight junction protein ZO-1 in hypoxic cochlear strial marginal cells in neonatal rats

In the cochlea, extracellular ATP (eATP) plays an important role in both physiological and pathological processes, but its role in the hypoxic cochlea remains unclear. The present study aims to investigate the relationship between eATP and hypoxic marginal cells (MCs) in the stria vascularis in cochlea. Combining various methodologies, we found that eATP accelerates cell death and decreases tight junction protein zonula occludens-1 (ZO-1) in hypoxic MCs. Flow cytometry and western blot analyses revealed an increase in apoptosis levels and suppression of autophagy, indicating that eATP causes additional cell death by increasing the apoptosis of hypoxic MCs. Given that autophagy inhibits apoptosis to protect MCs under hypoxia, apoptosis is probably enchanced by suppressing autophagy. Interleukin-33(IL-33)/suppression of tumorigenicity-2(ST-2)/matrix metalloprotein 9(MMP9) pathway activation was also observed during the process. Further experiments involving the use of additional IL-33 protein and an MMP9 inhibitor indicated that this pathway is responsible for the damage to the ZO-1 protein in hypoxic MCs. Our study revealed an adverse effect of eATP on the survival and ZO-1 protein expression of hypoxic MCs, as well as the underlying mechanism.

Tuberous sclerosis complex-1 (TSC1) contributes to selective neuronal vulnerability in Alzheimer's disease.

Selective neuronal vulnerability of hippocampal Cornu Ammonis (CA)-1 neurons is a pathological hallmark of Alzheimer's disease (AD) with an unknown underlying mechanism. We interrogated the expression of tuberous sclerosis complex-1 (TSC1; hamartin) and mTOR-related proteins in hippocampal CA1 and CA3 subfields. A human post-mortem cohort of mild (n= 7) and severe (n= 10) AD and non-neurological controls (n= 9) was used for quantitative and semi-quantitative analyses. We also developed an in vitro TSC1 knockdown model in rat hippocampal neurons, and transcriptomic analyses of TSC1 knockdown neuronal cultures were performed. We found a selective increase of TSC1 cytoplasmic inclusions in human AD CA1 neurons with hyperactivation of one of TSC1's downstream targets, the mammalian target of rapamycin complex-1 (mTORC1), suggesting that TSC1 is no longer active in AD. TSC1 knockdown experiments showed accelerated cell death independent of amyloid-beta toxicity. Transcriptomic analyses of TSC1 knockdown neuronal cultures revealed signatures that were significantly enriched for AD-related pathways. Our combined data point to TSC1 dysregulation as a key driver of selective neuronal vulnerability in the AD hippocampus. Future work aimed at identifying targets amenable to therapeutic manipulation is urgently needed to halt selective neurodegeneration, and by extension, debilitating cognitive impairment characteristic of AD.

Phased grapevine genome sequence of an Rpv12 carrier for biotechnological exploration of resistance to Plasmopara viticola.

The downy mildew disease caused by the oomycete Plasmopara viticola is a serious threat for grapevine and can cause enormous yield losses in viticulture. The quantitative trait locus Rpv12, mediating resistance against P. viticola, was originally found in Asian Vitis amurensis. This locus and its genes were analyzed here in detail. A haplotype-separated genome sequence of the diploid Rpv12-carrier Gf.99-03 was created and annotated. The defense response against P. viticola was investigated in an infection time-course RNA-seq experiment, revealing approximately 600 upregulated Vitis genes during host-pathogen interaction. The Rpv12 regions of the resistance and the sensitivity encoding Gf.99-03 haplotype were structurally and functionally compared with each other. Two different clusters of resistance-related genes were identified within the Rpv12 locus. One cluster carries a set of four differentially expressed genes with three ACCELERATED CELL DEATH 6-like genes. The other cluster carries a set of six resistance gene analogs related to qualitative pathogen resistance. The Rpv12 locus and its candidate genes for P. viticola resistance provide a precious genetic resource for P. viticola resistance breeding. Newly developed co-segregating simple sequence repeat markers in close proximity to the R-genes enable its improved applicability in marker-assisted grapevine breeding.

Short Telomere Syndrome presenting with pulmonary fibrosis, liver cirrhosis and hepatopulmonary syndrome: a case report

BackgroundIdiopathic pulmonary fibrosis is thought to result from aberrant post-injury activation of epithelial cells leading to fibroblast proliferation and activation. A number of genetic aetiologies have been implicated in this disease process, including, among others, the short telomere syndromes. Short telomere syndromes follow an autosomal dominant pattern of inheritance resulting in shortened telomere length, which consequently leads to accelerated cell death. Organs with rapid cell turnover are most affected.Case presentationWe describe a case of a 53-year-old man with a chief complaint of cough and dyspnea on exertion. His presentation was otherwise significant for features of accelerated aging, including a history of osteoporosis and early greying, and a family history of pulmonary fibrosis in his father. Pulmonary function testing revealed a restrictive pattern with severely reduced diffusion capacity and high resolution CT of the chest showed diffuse lung disease with mild fibrosis, in pattern suggesting an alternative diagnosis to IPF. Biopsy of the lung was in keeping with chronic fibrosing interstitial pneumonia. Imaging of the abdomen showed splenomegaly, hepatic cirrhosis and portal hypertension. Transthoracic contrast echocardiogram showed intrapulmonary shunting consistent with hepatopulmonary syndrome. Given the constellation of early aging, idiopathic pulmonary fibrosis, cryptogenic cirrhosis and a family history of pulmonary fibrosis in this patient, the Short Telomere Syndrome was suspected. Peripheral blood was sent for Flow-cytometry FISH, which demonstrated granulocyte telomere length below the 10th percentile for the patient’s age, consistent with a diagnosis of Short Telomere Syndrome in this clinical context. Targeted genetic testing of mutations known to be associated with short telomere was negative though it was acknowledged that the full spectrum of disease-causing mutations remains unknown. Given the extensive fibrosis on biopsy and his progressive hypoxemia he was treated with mycophenolate and prednisone. Ultimately, he developed progressive respiratory failure and underwent double lung and concurrent liver transplant 18 months after the initial diagnosis was made.ConclusionsShort Telomere Syndrome is a rare cause of end stage organ disease and testing lacks sensitivity making diagnosis challenging. Organ transplant is still the mainstay of treatment. Nevertheless, disease identification is important because of implications for family member screening and the possibility of future treatment options.