Density-dependent Resistance Research Articles

Induction of stearoyl-CoA desaturase confers cell density-dependent ferroptosis resistance in melanoma.

Ferroptosis is a form of regulated cell death that is induced by inhibiting glutathione peroxidase 4 (GPX4), which eliminates lipid peroxidation. Ferroptosis induction is influenced by the cell environment. However, the cellular states altering ferroptosis susceptibility remain largely unknown. We found that melanoma cell lines became resistant to ferroptosis as cell density increased. Comparative transcriptome and metabolome analyses revealed that cell density-dependent ferroptosis resistance was coupled with a shift toward a lipogenic phenotype accompanied by strong induction of stearoyl-CoA desaturase (SCD). Database analysis of gene dependency across hundreds of cancer cell lines uncovered a negative correlation between GPX4 and SCD dependency. Importantly, SCD inhibition, either pharmacologically or through genetic knockout, sensitized melanoma cells to GPX4 inhibition, thereby attenuating ferroptosis resistance in cells at high density. Our findings indicate that transition to an SCD-inducing, lipogenic cell state produces density-dependent resistance to ferroptosis, which may provide a therapeutic strategy against melanoma.

Abstract 1384: Stearoyl-CoA desaturase confers cell density-dependent resistance to ferroptosis induced by inhibition of glutathione peroxidase 4 in melanoma cells

Abstract Ferroptosis, one of the programmed cell deaths, is induced by accumulation of lipid peroxidation. To protect from ferroptosis, cells possess a wide range of reduction systems including glutathione peroxidase 4 (GPX4), which reduces lipid peroxides to non-toxic lipid alcohols. Indeed, inhibiting GPX4 leads to ferroptosis in various cancer cells, but cellular sensitivity to GPX4 inhibition often varies depending on cell conditions. Some previous studies reported that an increase in cell density induces resistance to GPX4 inhibition, but the mechanisms of cell density-dependent resistance are not fully understood. In this study, we have also found that several melanoma cell lines become resistant to ferroptosis induced by GPX4 inhibition in a density-dependent manner. For example, a selective GPX4 inhibitor, RSL3, immediately induced cell death at low cell density of A375 within 6-8 hours, but did not at high density even after 24 hours. The acute cell death at low density was rescued by ferroptosis inhibitors (lipid peroxidation inhibitors or iron chelator), but not those of apoptosis and necrosis, supporting that this type of cell death is ferroptosis. To elucidate the mechanisms of ferroptosis resistance at high density, we first conducted transcriptome and metabolome analyses using the difference between low and high densities of A375. The gene ontology analysis on upregulated genes at high density indicated an increase in fatty acid metabolism. In contrast, nutritional stress was not implied from the transcriptional analysis, resulting from frequent changes of culture medium to avoid nutrition starvation. The enrichment analysis based on upregulated metabolites at high density similarly suggested increased de novo lipogenesis. Particularly, gene and protein levels of stearoyl-CoA desaturase (SCD) were induced in a density-dependent manner. Similar induction of SCD was observed in several melanoma cell lines. We further examined the role of SCD in density-dependent ferroptosis resistance. With this aim, pharmacological inhibition or genetic knockout of SCD was conducted on A375 at different densities (low and high) treated with RSL3. The result showed that SCD inhibition reduced RSL3 resistance at high density while having no effect on RSL3 sensitivity at low density. In summary, we found that SCD is upregulated in melanoma cells at high density and protects cells from ferroptosis induced by GPX4 inhibition. It is known that metastasizing melanomas in the blood are sensitive to ferroptosis, resembling cells at low density. This report might provide an explanation for the ferroptosis sensitivity of metastasizing melanomas and a strategy for ferroptosis-based therapy. Citation Format: Hitomi Shirahama, Yuri Tani, Satomi Tsukahara, Yuka Okamoto, Akiko Hasebe, Shingo Dan, Akihiro Tomida. Stearoyl-CoA desaturase confers cell density-dependent resistance to ferroptosis induced by inhibition of glutathione peroxidase 4 in melanoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1384.

Density-dependent resistance protects Legionella pneumophila from its own antimicrobial metabolite, HGA.

To persist in microbial communities, the bacterial pathogen Legionella pneumophila must withstand competition from neighboring bacteria. Here, we find that L. pneumophila can antagonize the growth of other Legionella species using a secreted inhibitor: HGA (homogentisic acid). Unexpectedly, L. pneumophila can itself be inhibited by HGA secreted from neighboring, isogenic strains. Our genetic approaches further identify lpg1681 as a gene that modulates L. pneumophila susceptibility to HGA. We find that L. pneumophila sensitivity to HGA is density-dependent and cell intrinsic. Resistance is not mediated by the stringent response nor the previously described Legionella quorum-sensing pathway. Instead, L. pneumophila cells secrete HGA only when they are conditionally HGA-resistant, which allows these bacteria to produce a potentially self-toxic molecule while restricting the opportunity for self-harm. We propose that established Legionella communities may deploy molecules such as HGA as an unusual public good that can protect against invasion by low-density competitors.

Instability growth for magnetized liner inertial fusion seeded by electro-thermal, electro-choric, and material strength effects

A critical limitation of magnetically imploded systems such as magnetized liner inertial fusion (MagLIF) [Slutz et al., Phys. Plasmas 17, 056303 (2010)] is the magneto-Rayleigh-Taylor (MRT) instability which primarily disrupts the outer surface of the liner. MagLIF-relevant experiments have showed large amplitude multi-mode MRT instability growth growing from surface roughness [McBride et al., Phys. Rev. Lett. 109, 135004 (2012)], which is only reproduced by 3D simulations using our MHD code Gorgon when an artificially azimuthally correlated initialisation is added. We have shown that the missing azimuthal correlation could be provided by a combination of the electro-thermal instability (ETI) and an “electro-choric” instability (ECI); describing, respectively, the tendency of current to correlate azimuthally early in time due to temperature dependent Ohmic heating; and an amplification of the ETI driven by density dependent resistivity around vapourisation. We developed and implemented a material strength model in Gorgon to improve simulation of the solid phase of liner implosions which, when applied to simulations exhibiting the ETI and ECI, gave a significant increase in wavelength and amplitude. Full circumference simulations of the MRT instability provided a significant improvement on previous randomly initialised results and approached agreement with experiment.

Bistable dynamics of an insect–pathogen model

We consider a model for insect–pathogen interaction where the insect population is divided into two groups, one group susceptible to disease and other resistant to disease. An individual born susceptible to or resistant to disease depends on the local population levels at the start of each generation. Here we consider density-dependent models of transmission because we characterize diseases that spread through environmental propagules or through random contact among individuals. We consider the case where the fraction of resistant individuals increases as the total population increases. White and Wilson (Theor. Popul. Biol.56, 163 (1999)) have reported the results of density-dependent monotonic increase of resistance class by choosing a particular type of function. In this paper, we have chosen a class of monotonic density-dependent resistance functions and studied their effects on insect–pathogen dynamics. In particular, we have investigated the effects of different types of monotonic density-dependent resistance on the bistable nature of the model. Numerical simulation results are presented and interpreted.

Resistance of cancer cells to the TRAIL-induced apoptosis in confluent cultures

TRAIL (TNF-alpha Related Apoptosis-Inducing Ligand) is an attractive candidate for anticancer therapy. TRAIL selectively kills tumor cells, without damaging normal cells. It is known that cancer cells can acquire resistance to chemotherapeutic drugs, oxidative stress in high density culture. This phenomenon appears as a multi-cell resistance, cell adhesion-dependent resistance, or density-dependent resistance of tumor cells. However, it is unclear whether such resistance appears in TRAIL-induced apoptosis. We observed that the resistance to izTRAIL of all tumor cell lines used was considerably increased in confluent cultures. The increase in tumor cell resistance in dense populations is not related to their proliferative status. It was shown that the dissociation of calcium-dependent cell-cell contacts with EGTA did not suppress tumor cell resistance to izTRAIL in confluent cultures. This phenomenon of cancer cells resistance to TRAIL-induced apoptosis should be considered in the development of methods of anticancer therapy.

Diffusive charge transport in graphene on SiO 2

We review our recent work on the physical mechanisms limiting the mobility of graphene on SiO 2. We have used intentional addition of charged scattering impurities and systematic variation of the dielectric environment to differentiate the effects of charged impurities and short-range scatterers. The results show that charged impurities indeed lead to a conductivity linear in density ( σ ( n ) ∝ n ) in graphene, with a scattering magnitude that agrees quantitatively with theoretical estimates; increased dielectric screening reduces the scattering from charged impurities, but increases the scattering from short-range scatterers. We evaluate the effects of the corrugations (ripples) of graphene on SiO 2 on transport by measuring the height–height correlation function. The results show that the corrugations cannot mimic long-range (charged impurity) scattering effects, and have too small an amplitude-to-wavelength ratio to significantly affect the observed mobility via short-range scattering. Temperature-dependent measurements show that longitudinal acoustic phonons in graphene produce a resistivity that is linear in temperature and independent of carrier density; at higher temperatures, polar optical phonons of the SiO 2 substrate give rise to an activated, carrier density-dependent resistivity. Together the results paint a complete picture of charge carrier transport in graphene on SiO 2 in the diffusive regime.

Density-dependent resistance of the gypsy moth Lymantria dispar to its nucleopolyhedrovirus, and the consequences for population dynamics

The processes controlling disease resistance can strongly influence the population dynamics of insect outbreaks. Evidence that disease resistance is density-dependent is accumulating, but the exact form of this relationship is highly variable from species to species. It has been hypothesized that insects experiencing high population densities might allocate more energy to disease resistance than those at lower densities, because they are more likely to encounter density-dependent pathogens. In contrast, the increased stress of high-density conditions might leave insects more vulnerable to disease. Both scenarios have been reported for various outbreak Lepidoptera in the literature. We tested the relationship between larval density and disease resistance with the gypsy moth (Lymantria dispar) and one of its most important density-dependent mortality factors, the nucleopolyhedrovirus (NPV) LdMNPV, in a series of bioassays. Larvae were reared in groups at different densities, fed the virus individually, and then reared individually to evaluate response to infection. In this system, resistance to the virus decreased with increasing larval density. Similarly, time to death was faster at high densities than at lower densities. Implications of density-resistance relationships for insect-pathogen population dynamics were explored in a mathematical model. In general, an inverse relationship between rearing density and disease resistance has a stabilizing effect on population dynamics.

Modelling Density-Dependent Resistance in Insect–Pathogen Interactions

We consider a mathematical model for a host–pathogen interaction where the host population is split into two categories: those susceptible to disease and those resistant to disease. Since the model was motivated by studies on insect populations, we consider a discrete-time model to reflect the discrete generations which are common among insect species. Whether an individual is born susceptible or resistant to disease depends on the local population levels at the start of each generation. In particular, we are interested in the case where the fraction of resistant individuals in the population increases as the total population increases. This may be seen as a positive feedback mechanism since disease is the only population control imposed upon the system. Moreover, it reflects recent experimental observations from noctuid moth–baculovirus interactions that pathogen resistance may increase with larval density. We find that the inclusion of a resistant class can stabilise unstable host–pathogen interactions but there is greatest regulation when the fraction born resistant is density independent. Nonetheless, inclusion of density dependence can still allow intrinsically unstable host–pathogen dynamics to be stabilised provided that this effect is sufficiently small. Moreover, inclusion of density-dependent resistance to disease allows the system to give rise to bistable dynamics in which the final outcome is dictated by the initial conditions for the model system. This has implications for the management of agricultural pests using biocontrol agents—in particular, it is suggested that the propensity for density-dependent resistance be determined prior to such a biocontrol attempt in order to be sure that this will result in the prevention of pest outbreaks, rather than their facilitation. Finally we consider how the cost of resistance to disease affects model outcomes and discover that when there is no cost to resistance, the model predicts stable periodic outbreaks of the insect population. The results are interpreted ecologically and future avenues for research to address the shortfalls in the present model system are discussed.

Density-dependent resistance to apoptosis in retinal cells

PURPOSE. To study effects of cell density on retinal cell survival. METHODS. Apoptotic cell death was induced in cultured retinal cells seeded at higher or lower density by various stimuli including simulated ischemia, excitotoxicity and antibody against heat shock protein 27 (hsp27). Quantitative analysis of apoptotic cells was performed using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling technique and flow cytometry. Cytoskeleton was examined using immunocytochemistry and specific staining of actin by phalloidin and DNase I. In addition, alterations in the cytoskeletal proteins, bcl-2 family of proteins and hsp27 were studied using western blotting. RESULTS. Incubation of the cells under apoptotic stimuli caused higher rates of apoptosis in lower density cultures as determined by TUNEL technique and flow cytometric analysis. Both morphologic examination of cytoskeleton and western blotting revealed that after incubation with various stressors, degradation of actin and tubulin was more prominent in lower density cultures compared to higher density cultures. The expression of bcl-2 and bcl-xL was higher and the expression of bax was lower in lower density cultures compared to higher density cultures at basal condition. After incubation with stressors, bcl-2 and bcl-xL expressions decreased and bax expression increased in both lower and higher density cultures. However, we observed that the expression of hsp27 was higher in higher density cultures than in lower density cultures in the presence or absence of apoptotic stimuli. CONCLUSIONS. These findings demonstrate that retinal cells are more resistant to apoptosis in higher density cultures, independent of the inducer. This might be partly due to protective activity of endogenous hsp27 in the cells at higher density, which contributes to cytoskeletal integrity in response to apoptotic stimuli.

High cell density-dependent resistance and P-glycoprotein-mediated multidrug resistance in mitoxantrone-selected chinese hamster cells

Mitoxantrone (MIT) resistance has been studied in a colony selected from the CHO AA8 parental line in one step under a low degree of selective pressure (9 nM). The cells of the clonal isolate AA8/MIT C1(0) were sensitive to 9 nM MIT at low cell density but able to grow at high density. Parental AA8 cells were not able to grow under the latter condition. Decreased MIT accumulation (−20%) was observed at this step (step 0) in the absence of overexpression of mdr RNA coding for the drug efflux pump P-glycoprotein. Furthermore, AA8/MIT C1(0) did not exhibit cross resistance to vincristine, Adriaroycin and etoposide at low cell density. During subsequent controlled growth for 2 months at high cell density in the presence of 9 nM drug, an additional selection occurred leading to a 4-fold MIT-resistant subline AA8/MIT C1(+). This subline was characterized at this step (step I) and after an additional 4 months of culture in the presence of 9 nM MIT (step II). Analysis of mdr gene expression and gene copy number showed an increase in mdr RNA and a pattern of mdr gene amplification which changed between step I and II. AA8/MIT C1(+)II exhibited a classical multidrug resistance phenotype with decreased accumulation of [ 14CIMIT and cross-resistance to vincristine, Adriamycin and etoposide. The ability to form the cleavable complex in the presence of etoposide in DNA topoisomerase II-containing nuclear extracts was identical in AA8/MIT C1(+)II and AA8 cell lines. These results demonstrate a new sequence of events in MIT resistance: low level of drug resistance at high cell density followed by mdr gene amplification.

Airways impedance during single breaths of foreign gases

The changes in airways resistance (Raw) and inertance (Iaw) during single inspirations of pure methane, helium, neon, and ethane at a flow of 0.1 l/s were measured in six healthy subjects by use of a forced-oscillation technique. Raw and Iaw were computed from respiratory transfer impedance obtained at a frequency of 20 Hz by applying pressure oscillations at the chest and measuring flow at the mouth with a bag-in-box system. Compared with the air data, the changes of Iaw after inhalation of 500 ml of gas averaged -41.1% with methane, -82.8% with helium, -25.8% with neon, and +4.8% with ethane. These changes were slightly less than the changes in gas density (-45%, -86%, -31%, and +5%, respectively). The inhaled volumes at which 50% of the changes had occurred (V50) did not differ significantly among gases and were approximately 100 ml. For Raw the data were more noisy than for Iaw; they were discarded in two subjects because of a strong and irreproducible volume dependence in air. Consistent differences were seen between the remaining subjects, one of whom exhibited a predominant viscosity dependence of Raw, one a predominant density dependence, and two an intermediate pattern. V50s were larger for Raw than for Iaw, indicating a more peripheral distribution of Raw. For Raw, V50s were lower with helium than with methane, in agreement with the notion that density-dependent resistance is located mainly in the large airways. The results suggest that some information on the serial distribution of Raw and Iaw may be derived from impedance measurements with foreign gases.

Density-dependent resistance to puromycin in cell cultures.

AbstractPig kidney cells exhibit resistance to puromycin when cultured at high cell densities, growing progressively in otherwise lethal concentrations of drug. Comparison of population growth of cultures inoculated at different densities indicates that survival in puromycin is a function of cell density. Uptake of puromycin from culture media is less at higher cell densities resulting in decreased inhibition of protein synthesis by puromycin. Enhanced survival in puromycin at high population densities is attributed to reduced accumulation of puromycin from the culture media.

Possible Explanation of the Current Density-Dependent Resistivity of Exploding Wires

As a result of current density-dependent resistivity in exploding wires, there is a corresponding variation of the total energy input to the wire. The behavior of gold wire in the resistance regions 4 x 10/sup 7/ and 1.2 x 10/sup 8/ amp/cm/sup 2/ was studied with the aid of Kerr-cell photographs. Breakdown modes in the formation of shunting air arcs and in the vaporized wire material itself were observed. (C.J.G.)