2C Cells Research Articles

Thermal Performance Enhancement of Lithium-Ion Batteries Using Phase Change Material and Fin Geometry Modification

The rapid increase in emissions and the depletion of fossil fuels have led to a rapid rise in the electric vehicle (EV) industry. Electric vehicles predominantly rely on lithium-ion batteries (LIBs) to power their electric motors. However, the charging and discharging processes of LIB packs generate heat, resulting in a significant decline in the battery performance of EVs. Consequently, there is a pressing need for effective battery thermal management systems (BTMSs) for lithium-ion batteries in EVs. In the current study, a novel experimental BTMS was developed for the thermal performance enhancement of an LIB pack comprising 2 × 2 cells. Three distinct fin configurations (circular, rectangular, and tapered) were integrated for the outer wall of the lithium-ion cells. Additionally, the cells were fully submerged in phase change material (PCM). The study considered 1C, 2C, and 3C cell discharge rates, affiliated with their corresponding volumetric heat generation rates. The combination of rectangular fins and PCM manifested superior performance, reducing the mean cell temperature by 29.71% and 28.36% compared to unfinned lithium-ion cells under ambient conditions at the 1C and 2C discharge rates. Furthermore, at the 3C discharge rate, lithium-ion cells equipped with rectangular fins demonstrated a delay of 40 min in reaching the maximum surface temperature of 40 °C compared to the unfinned ambient case. After 60 min of battery discharge at the 3C rate, the cell surface temperature of the rectangular fin case only reached 42.7 °C. Furthermore, numerical simulations showed that the Nusselt numbers for lithium-ion cells with rectangular fins improved by 9.72% compared to unfinned configurations at the 3C discharge rate.

Polyploid cell dynamics and death before and after PEG-treatment of a NIH/3T3 derived culture: vinblastine effects on the regulation of cell subpopulations heterogeneity

BackgroundNeoplastic subpopulations can include polyploid cells that can be involved in tumor evolution and recurrence. Their origin can be traced back to the tumor microenvironment or chemotherapeutic treatment, which can alter cell division or favor cell fusion, generating multinucleated cells. Their progeny, frequently genetically unstable, can result in new aggressive and more resistant to chemotherapy subpopulations. In our work, we used NIHs cells, previously derived from the NIH/3T3 line after serum deprivation, that induced a polyploidization increase with the appearance of cells with DNA content ranging from 4 to 24c. This study aimed to analyze the cellular dynamics of NIHs culture subpopulations before and after treatment with the fusogenic agent polyethylene glycol (PEG), which allowed us to obtain new giant polyploid cells. Successively, PEG-untreated and PEG-treated cultures were incubated with the antimicrotubular poison vinblastine. The dynamics of appearance, decrease and loss of cell subpopulations were evaluated by correlating cell DNA content to mono-multinuclearity resulting from cell fusion and division process alteration and to the peculiarities of cell death events.ResultsDNA microfluorimetry and morphological techniques (phase contrast, fluorescence and TEM microscopies) indicated that PEG treatment induced a 4–24c cell increase and the appearance of new giant elements (64–140c DNA content). Ultrastructural analysis and autophagosomal–lysosomal compartment fluorochromization, which allowed us to correlate cytoplasmic changes to death events, indicated that cell depletion occurred through distinct mechanisms: apoptotic death involved 2c, 4c and 8c cells, while autophagic-like death involved intermediate 12–24c cells, showing nuclear (lobulation/micronucleation) and autophagic cytoplasm alterations. Death, spontaneously occurring, especially in intermediate-sized cells, was increased after vinblastine treatment. No evident cell loss by death events was detected in the 64–140c range.ConclusionsPEG-treated NIHs cultures can represent a model of heterogeneous subpopulations originating from cell fusion and division process anomalies. Altogether, our results suggest that the different cell dynamics of NIHs subpopulations can affect the variability of responses to stimuli able to induce cell degeneration and death. Apoptptic, autophagic or hybrid forms of cell death can also depend on the DNA content and ability to progress through the cell cycle, which may influence the persistence and fate of polyploid cell descendants, also concerning chemotherapeutic agent action.

Phosphorus deficiency induces sexual reproduction in the dinoflagellate Prorocentrum cordatum

Nitrogen (N) and phosphorus (P) are essential elements whose availability promotes successful growth of phytoplankton and governs aquatic primary productivity. In this study, we investigated the effect of N and/or P deficiency on the sexual reproduction of Prorocentrum cordatum, the dinoflagellate with the haplontic life cycle which causes harmful algal blooms worldwide. In P. cordatum cultures, N and the combined N and P deficiency led to the arrest of the cell cycle in the G0/G1 phases and attenuation of cell culture growth. We observed, that P, but not N deficiency triggered the transition in the life cycle of P. cordatum from vegetative to the sexual stage. This resulted in a sharp increase in percentage of cells with relative nuclear DNA content 2C (zygotes) and the appearance of cells with relative nuclear DNA content 4C (dividing zygotes). Subsequent supplementation with phosphate stimulated meiosis and led to a noticeable increase in the 4C cell number (dividing zygotes). Additionally, we performed transcriptomic data analysis and identified putative phosphate transporters and enzymes involved in the phosphate uptake and regulation of its metabolism by P. cordatum. These include high- and low-affinity inorganic phosphate transporters, atypical alkaline phosphatase, purple acid phosphatases and SPX domain-containing proteins.

Characterization of Transcriptome Dynamics during Early Fruit Development in Olive (Olea europaea L.).

In the olive (Olea europaea L.), an economically leading oil crop worldwide, fruit size and yield are determined by the early stages of fruit development. However, few detailed analyses of this stage of fruit development are available. This study offers an extensive characterization of the various processes involved in early olive fruit growth (cell division, cell cycle regulation, and cell expansion). For this, cytological, hormonal, and transcriptional changes characterizing the phases of early fruit development were analyzed in olive fruit of the cv. 'Picual'. First, the surface area and mitotic activity (by flow cytometry) of fruit cells were investigated during early olive fruit development, from 0 to 42 days post-anthesis (DPA). The results demonstrate that the cell division phase extends up to 21 DPA, during which the maximal proportion of 4C cells in olive fruits was reached at 14 DPA, indicating that intensive cell division was activated in olive fruits at that time. Subsequently, fruit cell expansion lasted as long as 3 weeks more before endocarp lignification. Finally, the molecular mechanisms controlling the early fruit development were investigated by analyzing the transcriptome of olive flowers at anthesis (fruit set) as well as olive fruits at 14 DPA (cell division phase) and at 28 DPA (cell expansion phase). Sequential induction of the cell cycle regulating genes is associated with the upregulation of genes involved in cell wall remodeling and ion fluxes, and with a shift in plant hormone metabolism and signaling genes during early olive fruit development. This occurs together with transcriptional activity of subtilisin-like protease proteins together with transcription factors potentially involved in early fruit growth signaling. This gene expression profile, together with hormonal regulators, offers new insights for understanding the processes that regulate cell division and expansion, and ultimately fruit yield and olive size.

Rhizobial infection of 4C cells triggers their endoreduplication during symbiotic nodule development in soybean.

Symbiosis between legumes and rhizobia results in the formation of nitrogen-fixing root nodules. Endoreduplication is essential for nodule development and efficient nitrogen fixation; however, the cellular mechanism by which rhizobial infection causes endoreduplication in symbiotic nodules and the roles of the resulting polyploid cells in nitrogen fixation remain largely unknown. Here, we developed a series of different approaches to separate infected cells (ICs) and uninfected cells (UCs) and determined their ploidy levels in soybean (Glycine max) developing nodules. We demonstrated that 4C nuclei exist in both UCs and ICs of developing nodules and that these 4C cells are primarily invaded by rhizobia and subsequently undergo endoreduplication. Furthermore, RNA-sequencing analysis of nuclei with different ploidy levels from soybean nodules at 12 d post-infection (dpi) and 20 dpi showed that 4C cells are predominantly ICs in 12-dpi nodules but UCs in 20-dpi nodules. We conclude that the infection of 4C cells by rhizobia is critical for initiating endoreduplication. These findings provide significant insight into rhizobial infection, nodule endoreduplication and nitrogen fixation in symbiotic nodules.

Planarian stem cells specify fate yet retain potency during the cell cycle.

Planarian whole-body regeneration is enabled by stem cells called neoblasts. At least some neoblasts are individually pluripotent. Neoblasts are also heterogeneous, with subpopulations of specialized neoblasts having different specified fates. Fate specification in neoblasts is regulated by fate-specific transcription factor (FSTF) expression. Here, we find that FSTF expression is common in neoblast S/G2/M cell-cycle phases but less common in G1. We find that specialized neoblasts can divide to produce progeny with asymmetric cell fates, suggesting that they could retain pluripotency. Furthermore, no known neoblast class was present in all neoblast colonies, suggesting that pluripotency is not the exclusive property of any known class. We tested this possibility with single-cell transplantations, which indicate that at least some specialized neoblasts are likely clonogenic. On the basis of these findings, we propose a model for neoblast pluripotency in which neoblasts can undergo specialization during the cell cycle without loss of potency.

Study of Hepatocytes Polyploidization Peculiarities in Cholestatic Liver of Adult Rats

According to the literature, different mechanisms and kinetics proceeding of regenerative growth has been established using the basic models of liver regeneration (after resection or chemically induced). Hence, in order to determine general regularities of the adaptive-compensatory processes in various pathological conditions, the processes taking place in the cholestatic liver of adult white rats during the first four days after common bile duct ligation have been studied. It has been shown that in cholestatic liver, compensatory-adaptive processes take place with different kinetics compared to those after resection. In particular, in response to the increased functional load caused by destructive processes during cholestasis, the liver, at an early stage, responds by simple division of high ploidy (binuclear tetraploid) cells and further provides their quantitative increase. The difference between the processes taking place in cholestatic and resected liver is more expressed on the third and fourth day after common bile duct ligation. In particular, 4c cells are still highest in cholestatic liver, while all ploidy cells are present in equal numbers in the regenerated liver after resection. This fact of compensatory growth characteristic for reparative regeneration was not detected in cholestatic liver at the mentioned date.

The Combination of the M2 Muscarinic Receptor Agonist and Chemotherapy Affects Drug Resistance in Neuroblastoma Cells.

One of the major limits of chemotherapy is depending on the ability of the cancer cells to elude and adapt to different drugs. Recently, we demonstrated how the activation of the M2 muscarinic receptor could impair neuroblastoma cell proliferation. In the present paper, we investigate the possible effects mediated by the preferential M2 receptor agonist arecaidine propargyl ester (APE) on drug resistance in two neuroblastoma cell lines, SK-N-BE and SK-N-BE(2C), a sub-clone presenting drug resistance. In both cell lines, we compare the expression of the M2 receptor and the effects mediated by the M2 agonist APE on cell cycle, demonstrating a decreased percentage of cells in S phase and an accumulation of SK-N-BE cells in G1 phase, while the APE treatment of SK-N-BE(2C) cells induced a block in G2/M phase. The withdrawal of the M2 agonist from the medium shows that only the SK-N-BE(2C) cells are able to rescue cell proliferation. Further, we demonstrate that the co-treatment of low doses of APE with doxorubicin or cisplatin significantly counteracts cell proliferation when compared with the single treatment. Analysis of the expression of ATP-binding cassette (ABC) efflux pumps demonstrates the ability of the M2 agonist to downregulate their expression and that this negative modulation may be dependent on N-MYC decreased expression induced by the M2 agonist. Our data demonstrate that the combined effect of low doses of conventional drugs and the M2 agonist may represent a new promising therapeutic approach in neuroblastoma treatment, in light of its significant impact on drug resistance and the possible reduction in the side effects caused by high doses of chemotherapy drugs.

Dysregulation of MITF Leads to Transformation in MC1R-Defective Melanocytes.

The MC1R/cAMP/MITF pathway is a key determinant for growth, differentiation, and survival of melanocytes and melanoma. MITF-M is the melanocyte-specific isoform of Microphthalmia-associated Transcription Factor (MITF) in human melanoma. Here we use two melanocyte cell lines to show that forced expression of hemagglutinin (HA) -tagged MITF-M through lentiviral transduction represents an oncogenic insult leading to consistent cell transformation of the immortalized melanocyte cell line Hermes 4C, being a melanocortin-1 receptor (MC1R) compound heterozygote, while not causing transformation of the MC1R wild type cell line Hermes 3C. The transformed HA-tagged MITF-M transduced Hermes 4C cells form colonies in soft agar and tumors in mice. Further, Hermes 4C cells display increased MITF chromatin binding, and transcriptional reprogramming consistent with an invasive melanoma phenotype. Mechanistically, forced expression of MITF-M drives the upregulation of the AXL tyrosine receptor kinase (AXL), with concomitant downregulation of phosphatase and tensin homolog (PTEN), leading to increased activation of the PI3K/AKT pathway. Treatment with AXL inhibitors reduces growth of the transformed cells by reverting AKT activation. In conclusion, we present a model system of melanoma development, driven by MITF-M in the context of MC1R loss of function, and independent of UV exposure. This model provides a basis for further studies of critical changes in the melanocyte transformation process.

Role of pyroglutamylated RFamide peptide43 in germ cell dynamics in mice testes in relation to energy metabolism

QRFP is a neuropeptide that regulates glucose homeostasis and increases insulin sensitivity in tissues. We have previously shown that QRFP and its receptor (GPR103) are predominantly expressed in germ cells and Sertoli cells, respectively, in mice testes. In the present study, we report that QRFP caused an increase in PCNA and a decrease in p27Kip1 expressions in the testis under both in vivo and ex vivo conditions. Besides, via an in vivo study, cell cycle analysis by FACS showed an increase in 2C cells and a decrease in 1C cells. QRFP also induced expression of GDNF and phosphorylation of Akt and ERK-1/2. Together these results suggest that QRFP has a proliferative effect on germ cells in mice testes, since it caused a proportional increase in the mitotic activity and the number of spermatogonial cells. Further, observations of increased expressions of STAT-3 and Neurog3 in treated mice suggest that QRFP treatment regulates priming of undifferentiated spermatogonia to undergo differentiation, while a decrease in c-Kit expression indicate that spermatogonia at this time point are in an undifferentiated state. In addition, QRFP administration also caused an increase in intratesticular levels of glucose and lactate, and in LDH activity accompanied by increased expressions of GLUT-3 and LDH-C in the testis. Also, the phosphorylation of IR-β and expressions of p-Akt and p-mTOR were increased under ex vivo conditions in testicular tissue. In conclusion, our findings suggest that QRFP treatment caused proliferation of germ cells independently from the hypothalamic-pituitary axis via regulation of testicular energy metabolism.

Calcium signaling mediated by aminergic GPCRs is impaired by the PI3K inhibitor LY294002 and its analog LY303511 in a PI3K-independent manner

The phosphoinositide 3-kinase (PI3K) inhibitor LY294002 (LY294) and its much less active analog LY303511 (LY303) constitute the paired probe that is commonly used to demonstrate the involvement of PI3K in intracellular signaling. We studied effects of LY294 and LY303 on Ca2+ signaling initiated by certain GPCR agonists in cells of several lines, including CHO cells expressing the recombinant serotonin receptor 5-HT2C and mesenchymal stromal cells derived from the human adipose tissue (AD-MSCs) and umbilical cord (UD-MSCs). The LY294/LY303 pair exerted apparently specific effects on responsiveness of AD-MSCs to ATP, suggesting the involvement of PI3K in ATP transduction. Surprisingly, LY303 inhibited Ca2+ transients elicited by histamine in the same cells, while LY294 was ineffective. This observation and other findings implicated a PI3K-unrelated mechanism in mediating effects of the LY compound on AD-MSC responsiveness to histamine. With LY303 in the bath, the dose dependence of histamine responses was shifted positively at the invariable number of responsive cells, as would be the case with a competitive antagonist of histamine receptors. Moreover, LY303 and LY294 inhibited Ca2+ transients elicited by acetylcholine and serotonin in UD-MSCs and CHO/5-HT2C cells, respectively. Our overall results argued for the possibility that LY294 and LY303 could directly affect activity of aminergic GPCRs. Thus, LY303511 and LY294002 should be used cautiously in studies of PI3K as a factor of GPCR signaling.

Arabidopsis nucleoporin CPR5 controls trichome cell death through the core cell cycle regulator CKI.

The Arabidopsis trichome is a polyploid epidermal cell resulting from multiple rounds of endocycles. The CYCLIN-DEPENDENT KINASE INHIBITOR (CKI) family proteins are core cell cycle regulators that promote the endocycle. CONSTITUTIVE EXPRESSION OF PR GENES 5 (CPR5) is a plant-specific nucleoporin. It has been found that two Arabidopsis CKI, SIAMESE (SIM) and SIAMESE-RELATED 1 (SMR1), function downstream of CPR5 to activate plant effector-triggered cell death. The sim smr1 double mutants form multicellular and clustered trichomes, while the cpr5 mutants produce dead and branchless trichomes. This study explored roles of the CPR5-CKI signalling pathway in trichome cell cycle transition. To examine the underlying mechanism of how cell cycle transition is regulated in plant trichomes, Trypan blue staining, flow cytometry, scanning electron microscopy (SEM) and nuclear DNA measurement were conducted. The native promoter-driven CKI and GUS fusion reporter showed that both SIM and SMR1 proteins were preferentially expressed in trichomes. The cpr5-induced dead and branchless trichomes were fully suppressed by the sim smr1 double mutant, suggesting that SIM and SMR1 function downstream of CPR5 in trichome development. Flow cytometry analysis showed that as compared to the number of 2C (C=DNA content in a haploid nucleus) cells, the number of 4C cells significantly increased, whereas that of polyploidy cells (8C and 16C) dramatically decreased in the cpr5 mutant. The elevated 4C/2C ratio in the cpr5 mutant is consistent with de-repression of pro-endocycle regulators SIM and SMR1. The polyploidy cells (8C and 16C) may be selectively targeted to cell death, which is therefore attributed to the branchless trichomes in the cpr5 mutant. Nuclear DNA content analysis demonstrated that the nuclear DNA content of trichomes in the cpr5 sim mutant was significantly higher than in the sim mutant, indicating that CPR5 is a negative endocycle regulator in trichomes. This study reveals that the CPR5-CKI signalling pathway controls trichome cell cycle transition and excessive endocycles are required for cell death in plant trichomes.

Abstract 1049A: Deregulation of MITF in the context of inactive MC1R leads to melanocyte transformation

Abstract MC1R, the cAMP pathway and the response to solar UV are all elements involved in the growth, differentiation and survival of melanocytes and in malignant melanoma. Here we present a model of melanomagenesis based on the forced expression of MITF-M in the context of MC1R-inactivating variants in a way not dependent on sun exposure. We demonstrate that lentiviral transduction of MITF-M alone leads to consistent transformation of the Hermes4C cell line with inactive MC1R, but not the Hermes3C cell line having a wild type background. Hermes 4c cells transduced with MITF-M are able to form tumors in mice. We mapped MITF binding to chromatin and found that 4C cells displayed enhanced MITF chromatin binding compared to 3C. Expression analysis revealed that the altered binding pattern leads to enhanced transcription of epithelial to mesenchymal transition genes and consequent repression of key melanocyte-specific genes e.g. involved in pigmentation in the background of inactive MC1R. We observed a marked deregulation of AXL and EGFR, which is accompanied by downregulation of PTEN and increased phosphorylation of ERK and PI3K-AKT pathway. In conclusion, we show that MITF-M is a molecular switch in the context of MC1R-inactivating variant which leads to cell reprogramming and melanomagenesis. Citation Format: Tine Norman Alver, Timothy J. Lavelle, Karen-Marie Heintz, Patrik Wernhoff, Vegard Nygaard, Geir F. Øy, Sigurd L. Bøe, Alfonso Urbanucci, Eivind J. Hovig. Deregulation of MITF in the context of inactive MC1R leads to melanocyte transformation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1049A.

Avenanthramide Aglycones and Glucosides in Oat Bran: Chemical Profile, Levels in Commercial Oat Products, and Cytotoxicity to Human Colon Cancer Cells.

Avenanthramides (AVAs), unique phytochemicals in oat, have attracted an increasing amount of attention due to their outstanding health benefits. However, the chemical profile and the levels of AVAs in commercial oat products as well as their health benefits have not been examined in detail. In the present study, a total of 29 AVA aglycones and AVA glucosides were identified and characterized from oat bran using NMR (1D and 2D NMR) and LC-MS techniques. Among them, 17 novel AVA glucosides were reported in oat bran for the first time. The most abundant AVA glucoside, 2c-3'- O-glc, had a similar growth inhibitory activity with the major AVA, 2c, against HCT-116 and HT-29 human colon cancer cells, indicating glucosylation does not affect the growth inhibitory effects of AVAs. Furthermore, the levels of all individual AVAs in 13 commercial oat products were analyzed using HPLC-MS/MS. The total AVAs contents in various oat products ranged from 9.22 to 61.77 mg/kg (fresh weight).

DNA double-strand breaks promote endoreduplication in radish cotyledon.

DSBs differently affect endoreduplication and organ size in radish cotyledons and hypocotyls in different light conditions, suggesting that DSBs-mediated endoreduplication varies based on different developmental and environmental cues. Endoreduplication induced by DNA double strand breaks (DSBs) in Arabidopsis thaliana roots and cultured cells has been reported in recent years. In this study, we investigated whether DSBs-mediated endoreduplication also occurs in other tissues, such as cotyledons and hypocotyls of radish (Raphanus sativus var. longipinnatus) plants. To induce DSBs, UV irradiation and Zeocin treatment were applied to in vitro-cultured radish seedlings, and ploidy distribution of the treated tissues was analyzed by flow cytometry. Consequently, frequencies of the higher ploidy (8C) cells and cycle values in the cotyledon tissues increased with increasing doses of UV irradiation and concentrations of Zeocin, irrespective of light conditions. UV-stimulated endoreduplication was also observed in four Brassica species. In hypocotyls, UV treatments decreased the frequencies of higher ploidy (32C) cells and cycle values in dark-grown seedlings, whereas Zeocin treatments increased the frequencies of higher ploidy (16C and 32C) cells and cycle values in light- and dark-grown seedlings. Among the treatments, organ sizes did not simply correlate with cycle values. The effects of treatments on endoreduplication and organ size differed based on organ and light conditions, indicating that DSBs-mediated endoreduplication may involve a multifaceted response to different developmental and environmental cues.

Genomic Instability in Cancer I: DNA-Repair Triggering Primitive Hereditary 4n-Skewed, Amitotic Division-System, the Culprit in EMT/MET/Metaplasia Cancer-Concepts

The objective was to gain proof of genome damage-repair induced mitotic slippage process (MSP) to 4n-diplochromosome skewed division-system, earlier suggested to have “cancer-deciding” consequences. Our damage-model showed two succeeding phases: molecular mutations for initiation of fitness-gained cells, and large chromosomal changes to aneuploidy from inherited DNA-breakage-repair inaccuracies. The mutations were gained while DNA-repair and DNA-replication, co-existed in the route to tetraploidy, a phenomenon also expressed for some existing unicellular organisms. These organisms also showed genome reductive, amitotic, meioticlike division, and was the origin of human genome conserved, self-inflicted 90° reorientation of the 4n nucleus relative to the cytoskeleton axis. In the in vitro DNA-damage model, this remarkable 4n-event deciding “flat-upright” cell-growth characteristics showed several consequences, for example, cancer-important, E-cadherin-β-catenin cell-to-cell adherence destruction, which gave diploid progeny cells, mobility freedom from cell contact inhibition, likely in renewal tissues. This 4n-skewed division-system with inheritance in progeny cells for repeat occurrences as mentioned for flat-up-right growth patterns is similar to claimed concepts of metaplasia-EMT/MET embryogenesis events in cancer evolution. A scrutiny of this literature, proof-wise invalidated this embryological concept by tetraploid 8C cells occurring in MET events and, was noted for small cell occurrence, i.e., diploidy from 4n-8C reductive division, an also event for tumor relapse cells, derived from genome damaging therapy agents. Pre-cancer hyperplasia reported MSP, cadherincatenin destruction and 90° perpendicularity to basal cell membrane. The DNA-damage-repair model can weed-out therapy-agents triggering 4n-skewed division. Cancer-control, beginning-information, is likely from mutational identity of the 4n derived fitness-gained cells.

Life-history stages of natural bloom populations and the bloom dynamics of a tropical Asian ribotype of Alexandrium minutum

In 2015, a remarkably high density bloom of Alexandrium minutum occurred in Sungai Geting, a semi-enclosed lagoon situated in the northeast of Peninsular Malaysia, causing severe discoloration and contaminated the benthic clams (Polymesoda). Plankton and water samples were collected to investigate the mechanisms of bloom development of this toxic species. Analysis of bloom samples using flow cytometry indicated that the bloom was initiated by the process of active excystment, as planomycetes (>4C cells) were observed in the early stage of the bloom. Increase in planozygotes (2C cells) was evident during the middle stage of the bloom, coinciding with an abrupt decrease in salinity and increase of temperature. The bloom was sustained through the combination of binary division of vegetative cells, division of planozygotes, and cyst germination through continuous excystment. Nutrient depletion followed by precipitation subsequently caused the bloom to terminate. This study provides the first continuous record of in situ life-cycle stages of a natural bloom population of A. minutum through a complete bloom cycle. The event has provided a fundamental understanding of the pelagic life-cycle stages of this tropical dinoflagellate, and demonstrated a unique bloom development characteristic shared among toxic Alexandrium species in coastal embayments.

Poly(ADP-ribose) polymerases regulate cell division and development in Arabidopsis roots.

Root organogenesis involves cell division, differentiation and expansion. The molecular mechanisms regulating root development are not fully understood. In this study, we identified poly(adenosine diphosphate (ADP)-ribose) polymerases (PARPs) as new players in root development. PARP catalyzes poly(ADP-ribosyl)ation of proteins by repeatedly adding ADP-ribose units onto proteins using nicotinamide adenine dinucleotide (NAD+ ) as the donor. We found that inhibition of PARP activities by 3-aminobenzomide (3-AB) increased the growth rates of both primary and lateral roots, leading to a more developed root system. The double mutant of Arabidopsis PARPs, parp1parp2, showed more rapid primary and lateral root growth. Cyclin genes regulating G1-to-S and G2-to-M transition were up-regulated upon treatment by 3-AB. The proportion of 2C cells increased while cells with higher DNA ploidy declined in the roots of treated plants, resulting in an enlarged root meristematic zone. The expression level of PARP2 was very low in the meristematic zone but high in the maturation zone, consistent with a role of PARP in inhibiting mitosis and promoting cell differentiation. Our results suggest that PARPs play an important role in root development by negatively regulating root cell division.

Transfer of Allogeneic CD4+ T Cells Rescues CD8+ T Cells in Anti-PD-L1-Resistant Tumors Leading to Tumor Eradication.

Adoptively transferred CD8+ T cells can stabilize the size of solid tumors over long periods of time by exclusively recognizing antigen cross-presented on tumor stroma. However, these tumors eventually escape T-cell-mediated growth control. The aim of this study was to eradicate such persistent cancers. In our model, the SIYRYYGL antigen is expressed by cancer cells that lack the MHC-I molecule Kb needed for direct presentation, but the antigen is picked up and cross-presented by tumor stroma. A single injection of antigen-specific 2C CD8+ T cells caused long-term inhibition of tumor growth, but without further intervention, tumors started to progress after approximately 3 months. Escape was associated with reduced numbers of circulating 2C cells. Tumor-infiltrating 2C cells produced significantly less TNFα and expressed more of the "exhaustion" markers PD-1 and Tim-3 than T cells from lymphoid organs. High-dose local ionizing radiation, depletion of myeloid-derived suppressor cells, infusions of additional 2C cells, and antibodies blocking PD-L1 did not prevent tumor escape. In contrast, adoptive transfer of allogeneic CD4+ T cells restored the numbers of circulating Ag-specific CD8+ T cells and their intratumoral function, resulting in tumor eradication. These CD4+ T cells had no antitumor effects in the absence of CD8+ T cells and recognized the alloantigen cross-presented on tumor stroma. CD4+ T cells might also be effective in cancer patients when PD-1/PD-L1 blockade does not rescue intratumoral CD8+ T-cell function and tumors persist. Cancer Immunol Res; 5(2); 127-36. ©2017 AACR.

Synthesis, structure-activity relationship studies and biological evaluation of novel 2,5-disubstituted indole derivatives as anticancer agents.

Three novel series of 2,5-disubstituted indole derivatives were synthesized and evaluated in vitro for their antiproliferative activity against human cancer cells and HIV-1 inhibition activity used as a readout of cellular activity. Most compounds were found to have potent anticancer activity. In particular, 2c and 3b which showed effectively to repress HIV-1 transcription had a pan antiproliferative activity in cervical cancer cells (HeLa), breast cancer cells (MCF-7), liver cancer cells (HepG2), and lung cancer cells (H460 and A549). While 3b exhibited high sensitivity to A549 cells with the IC50 value 0.48±0.15μm, 2c showed high selectivity toward HepG2 cells with the IC50 value 13.21±0.30μm. With respect to the cellular mechanism of action, HepG2 cells treated with 2c and A549 cells treated with 3b for 24h were studied by annexin V/PI staining and Western blot analysis, and results revealed that 2c and 3b may induce cancer cells apoptosis through inhibiting the phosphorylation at Ser2 of RNAPII CTD which can be phosphorylated by cyclin-dependent kinase 9. These studies indicated that 2c and 3b may develop as potent lead compounds in the therapy of cancer. However, determining their roles in preventing HIV-1 still requires further intensive study.