Transcription factor GATA4 and apoptotic TRAIL pathway in granulosa cell function and tumors

Normal growth and development require the precise control of gene expression. Transcription factors are proteins that regulate gene expression by binding specific sequences of DNA. Abnormalities in transcription are implicated in a variety of human diseases, including cancer, endocrine disorders and birth defects. Transcription factor GATA4 has emerged as an important regulator of normal development and function in a variety of endoderm- and mesoderm- derived tissues, including gut, heart and several endocrine organs, such as gonads. Mice harboring a null mutation of Gata4 gene die during embryogenesis due to failure in heart formation, complicating the study of functional role of GATA4 in other organs. However, the expression pattern of GATA4 suggests it may play a role in the regulation of ovarian granulosa cell development, function and apoptosis. This premise is supported by in vitro studies showing that GATA4 regulates several steroidogenic enzymes as well as auto-, para- and endocrine signaling molecules important for granulosa cell function. This study assessed the in vivo role of GATA4 for granulosa cell function by utilizing two genetically modified mouse strains. The findings in the GATA4 deficient mice included delayed puberty, impaired fertility and signs of diminished estrogen production. At the molecular level, the GATA4 deficiency leads to attenuated expression of central steroidogenic genes, Steroidogenic acute regulatory protein (StAR), Side-chain cleavage (SCC), and aromatase as a response to stimulations with exogenous gonadotropins. Taken together, these suggest GATA4 is necessary for the normal ovarian function and female fertility. Programmed cell death, apoptosis, is a crucial part of normal ovarian development and function. In addition, disturbances in apoptosis have been implicated to pathogenesis of human granulosa cell tumors (GCTs). Apoptosis is controlled by extrinsic and intrinsic pathways. The intrinsic pathway is regulated by members of Bcl-2 family, and its founding member, the anti-apoptotic Bcl-2, is known to be important for granulosa cell survival. This study showed that the expression levels of GATA4 and Bcl-2 correlate in the human GCTs and that GATA4 regulates Bcl-2 expression, presumably by directly binding to its promoter. In addition, disturbing GATA4 function was sufficient to induce apoptosis in cultured GCT- derived cell line. Taken together, these results suggest GATA4 functions as an anti-apoptotic factor in GCTs. The extrinsic apoptotic pathway is controlled by the members of tumor necrosis factor (TNF) superfamily. An interesting ligand of this family is TNF-related apoptosis-inducing ligand (TRAIL), possessing a unique ability to selectively induce apoptosis in malignant cells. This study characterized the previously unknown expression of TRAIL and its receptors in both developing and adult human ovary, as well as in malignant granulosa cell tumors. TRAIL pathway was shown to be active in GCTs suggesting it may be a useful tool in treating these malignancies. However, more studies are required to assess the function of TRAIL pathway in normal ovaries. In addition to its ability to induce apoptosis in GCTs, this study revealed that GATA4 protects these malignancies from TRAIL-induced apoptosis. GATA4 presumably exerts this effect by regulating the expression of anti-apoptotic Bcl-2. This is of particular interest as high expression of GATA4 is known to correlate to aggressive GCT behavior. Thus, GATA4 seems to protect GCTs from endogenous TRAIL by upregulating anti-apoptotic factors such as Bcl-2.

Interfacial coupling and its size dependence in PbTiO3 and PbMg1/3Nb2/3O3 multilayers

Multilayers of Pb(Mg1/3Nb2/3)O-3 (PMN)-PbTiO3 (PT) were deposited through pulsed laser ablation deposition with different periodicities (d=10, 20, 30, 40, 50, 60, and 70 nm) for a constant total thickness of the film. The presence of superlattice reflections in the x-ray diffraction pattern clearly showed the superlattice behavior of the fabricated structures over a periodicity range of 20-50 nm. Polarization hysteresis and the capacitance-voltage characteristics of these films show clear size dependent ferroelectric and antiferroelectric (AFE) characteristics. Presence of long-range coupling and strain in multilayers with lower periodicity (similar to 10 nm) exhibited a clear ferroelectric behavior similar to a solid solution of PMN and PT. Multilayers with higher periodicities (20-50 nm) exhibited antiferroelectric behavior, which could be understood from the energy arguments. On further increase of periodicity, they again exhibit ferroelectric behavior. The polarization studies were carried out beyond the Curie temperature T-c of PMN to understand the interlayer interaction. The interaction is changed to a ferroelectric-paraelectric interlayer and tends to lose its antiferroelectric behavior. The behavior of remnant polarization P-r and dP(r)/dT with temperature clearly proves that the AFE coupling of these superlattices is due to the extrinsic interfacial coupling and not an intrinsic interaction as in a homogeneous conventional AFE material. The evidence of an averaged behavior at a periodicity of similar to 10 nm, and the behavior of individual materials at larger periodicities were further confirmed through dielectric phase transition studies. The presence of AFE interfacial coupling was insignificant over the dielectric phase transition of the multilayers.

Ricin-membrane interaction: membrane penetration depth by fluorescence quenching and resonance energy transfer

The entry of the plant toxin ricin and its A- and B-subunits in model membranes in the presence as well as absence of monosialoganglioside (GM(1)) has been studied. Dioleoylphosphatidylcholine and 5-, 10-, and 12-doxyl- or 9,10-dibromophosphatidylcholines serve as quenchers of intrinsic tryptophan fluorescence of the proteins. The parallax method of Chattopadhyay and London [(1987) Biochemistry 26, 39-45] has been employed to measure the average membrane penetration depth of tryptophans of ricin and its B-chain and the actual depth of the sole Trp 211 in the A-chain. The results indicate that both of the chains as well as intact ricin penetrate the membrane deeply and the C-terminal end of the A-chain is well inside the bilayer, especially at pH 4.5. An extrinsic probe N-(iodoacetyl)-N'-(5-sulfo-1-naphthyl) ethylenediamine (I-AEDANS) has been attached to Cys 259 of the A-chain, and the kinetics of penetration has been followed by monitoring the increase in AEDANS fluorescence at 480 nm. The insertion follows first-order kinetics, and the rate constant is higher at a lower pH. The energy transfer distance analysis between Trp 211 and AEDANS points out that the conformation of the A-chain changes as it inserts into the membrane. CD studies indicate that the helicity of the proteins increases after penetration, which implies that some of the unordered structure in the native protein is converted to the ordered form during this process. Hydrophobic forces seem to be responsible for stabilizing a particular protein conformation inside the membrane.

SiO Maser Emission and the Intrinsic Properties of Mira Variables

This paper presents observations of SiO maser emission from 161 Mira variables distributed over a wide range of intrinsic parameters like spectral type, bolometric magnitude and amplitude of pulsation. The observations were made at 86.243 GHz, using the 10.4 m millimeter-wave telescope of the Raman Research Institute at Bangalore, India. These are the first observations made using this telescope. From these observations, we have established that the maser emission is restricted to Miras having mean spectral types between M6 and M10. The infrared period-luminosity relation for Mira variables is used to calculate their distances and hence estimate their maser luminosities from the observed fluxes. The maser luminosity is found to be correlated with the bolometric magnitude of the Mira variable. On an H-R diagram, the masing Mira variables are shown to lie in a region distinct from that for the non-masing ones.

Novel Levamisole Derivative Induces Extrinsic Pathway of Apoptosis in Cancer Cells and Inhibits Tumor Progression in Mice

Background: Levamisole, an imidazo(2,1-b) thiazole derivative, has been reported to be a potential antitumor agent. In the present study, we have investigated the mechanism of action of one of the recently identified analogues, 4a (2-benzyl-6-(4'-fluorophenyl)-5-thiocyanato-imidazo2,1-b]1,3,4]thi adiazole). Materials and Methods: ROS production and expression of various apoptotic proteins were measured following 4a treatment in leukemia cell lines. Tumor animal models were used to evaluate the effect of 4a in comparison with Levamisole on progression of breast adenocarcinoma and survival. Immunohistochemistry and western blotting studies were performed to understand the mechanism of 4a action both ex vivo and in vivo. Results: We have determined the IC50 value of 4a in many leukemic and breast cancer cell lines and found CEM cells most sensitive (IC50 5 mu M). Results showed that 4a treatment leads to the accumulation of ROS. Western blot analysis showed upregulation of pro-apoptotic proteins t-BID and BAX, upon treatment with 4a. Besides, dose-dependent activation of p53 along with FAS, FAS-L, and cleavage of CASPASE-8 suggest that it induces death receptor mediated apoptotic pathway in CEM cells. More importantly, we observed a reduction in tumor growth and significant increase in survival upon oral administration of 4a (20 mg/kg, six doses) in mice. In comparison, 4a was found to be more potent than its parental analogue Levamisole based on both ex vivo and in vivo studies. Further, immunohistochemistry and western blotting studies indicate that 4a treatment led to abrogation of tumor cell proliferation and activation of apoptosis by the extrinsic pathway even in animal models. Conclusion: Thus, our results suggest that 4a could be used as a potent chemotherapeutic agent.

Extracts of Strawberry Fruits Induce Intrinsic Pathway of Apoptosis in Breast Cancer Cells and Inhibits Tumor Progression in Mice

Background: The consumption of berry fruits, including strawberries, has been suggested to have beneficial effects against oxidative stress mediated diseases. Berries contain multiple phenolic compounds and secondary metabolites that contribute to their biological properties. Methodology/Principal Findings: Current study investigates the anticancer activity of the methanolic extract of strawberry (MESB) fruits in leukaemia (CEM) and breast cancer (T47D) cell lines ex vivo, and its cancer therapeutic and chemopreventive potential in mice models. Results of MTT, trypan blue and LDH assays suggested that MESB can induce cytotoxicity in cancer cells, irrespective of origin, in a concentration-and time-dependent manner. Treatment of mice bearing breast adenocarcinoma with MESB blocked the proliferation of tumor cells in a time-dependent manner and resulted in extended life span. Histological and immunohistochemical studies suggest that MESB treatment affected tumor cell proliferation by activating apoptosis and did not result in any side effects. Finally, we show that MESB can induce intrinsic pathway of apoptosis by activating p73 in breast cancer cells, when tumor suppressor gene p53 is mutated. Conclusions/Significance: The present study reveals that strawberry fruits possess both cancer preventive and therapeutic values and we discuss the mechanism by which it is achieved.

A Novel Anticancer Agent, 8-Methoxypyrimido4 `,5 `: 4,5]thieno(2,3-b) Quinoline-4(3H)-One Induces Neuro 2a Neuroblastoma Cell Death through p53-Dependent, Caspase-Dependent and Independent Apoptotic Pathways

Neuroblastoma is the most common cancer in infants and fourth most common cancer in children. Despite recent advances in cancer treatments, the prognosis of stage-IV neuroblastoma patients continues to be dismal which warrant new pharmacotherapy. A novel tetracyclic condensed quinoline compound, 8-methoxypyrimido 4 `,5 `: 4,5] thieno(2,3-b) quinoline-4(3H)-one (MPTQ) is a structural analogue of an anticancer drug ellipticine and has been reported to posses anticancer property. Study on MPTQ on neuroblastoma cells is very limited and mechanisms related to its cytotoxicity on neuroblastoma cells are completely unknown. Here, we evaluated the anticancer property of MPTQ on mouse neuro 2a and human SH-SY5Y neuroblastoma cells and investigated the mechanisms underlying MPTQ-mediated neuro 2a cell death. MPTQ-mediated neuro 2a and SH-SY5Y cell deaths were found to be dose and time dependent. Moreover, MPTQ induced cell death reached approximately 99.8% and 90% in neuro 2a and SH-SY5Y cells respectively. Nuclear oligonucleosomal DNA fragmentation and Terminal dUTP Nick End Labelling assays indicated MPTQ-mediated neuro 2a cell death involved apoptosis. MPTQ-mediated apoptosis is associated with increased phosphorylation of p53 at Ser15 and Ser20 which correlates with the hyperphosphorylation of Ataxia-Telangiectasia mutated protein (ATM). Immunocytochemical analysis demonstrated the increased level of Bax protein in MPTQ treated neuro 2a cells. MPTQ-mediated apoptosis is also associated with increased activation of caspase-9, -3 and -7 but not caspase-2 and -8. Furthermore, increased level of caspase-3 and cleaved Poly ( ADP Ribose) polymerase were observed in the nucleus of MPTQ treated neuro 2a cells, suggesting the involvement of caspase-dependent intrinsic but not extrinsic apoptotic pathway. Increased nuclear translocation of apoptosis inducing factor suggests additional involvement of caspase-independent apoptosis pathway in MPTQ treated neuro 2a cells. Collectively, MPTQ-induced neuro 2a cell death is mediated by ATM and p53 activation, and Bax-mediated activation of caspase-dependent and caspase-independent mitochondrial apoptosis pathways.

Membrane-based deformable mirror: intrinsic aberrations and alignment issues

A deformable mirror (DM) is an important component of an adaptive optics system. It is known that an on-axis spherical/parabolic optical component, placed at an angle to the incident beam introduces defocus as well as astigmatism in the image plane. Although the former can be compensated by changing the focal plane position, the latter cannot be removed by mere optical realignment. Since the DM is to be used to compensate a turbulence-induced curvature term in addition to other aberrations, it is necessary to determine the aberrations induced by such (curved DM surface) an optical element when placed at an angle (other than 0 deg) of incidence in the optical path. To this effect, we estimate to a first order the aberrations introduced by a DM as a function of the incidence angle and deformation of the DM surface. We record images using a simple setup in which the incident beam is reflected by a 37 channel micro-machined membrane deformable mirror for various angles of incidence. It is observed that astigmatism is a dominant aberration, which was determined by measuring the difference between the tangential and sagittal focal planes. We justify our results on the basis of theoretical simulations and discuss the feasibility of using such a system for adaptive optics considering a trade-off between wavefront correction and astigmatism due to deformation. (C) 2015 Optical Society of America

Influences of strain rate and temperature variation on material intrinsic length of plasticity strain gradient theory

Cowper-Symonds and Johnson-Cook dynamic constitutive relations are used to study the influence of both strain rate effect and temperature variation on the material intrinsic length scale in strain gradient plasticity. The material intrinsic length scale decreases with increasing strain rates, and this length scale increases with temperature.

Intrinsic correlation between fragility and bulk modulus in metallic glasses

A systematic study on the available data of 26 metallic glasses shows that there is an intrinsic correlation between fragility of a liquid and bulk modulus of its glass. The underlying physics can be rationalized within the formalism of potential energy landscape thermodynamics. It is surprising to find that the linear correlation between the fragility and the bulk-shear modulus ratio exists strictly at either absolute zero temperature or very high frequency. Further analyses indicate that a real flow event in bulk metallic glasses is shear dominant, and fragility is in inverse proportion to shear-induced bulk dilatation. Finally, extension of these findings to nonmetallic glasses is discussed.

Intrinsic correlation between dilatation and pressure sensitivity of plastic flow in metallic glasses

Taking shear-induced dilatation into consideration in shear transformation zone (STZ) operations, we derive a new yield criterion that reflects the pressure sensitivity in plastic flow in metallic glasses (MGs), which agrees well with experiments. Furthermore, an intrinsic theoretical correlation between the pressure sensitivity coefficient and the dilatation factor is revealed. It is found that the pressure sensitivity of plastic flow of MGs originates in the dilatation of microscale STZs.

Earthquakes, influenza and cycles of Indian kala-azar.

It is suggested that previous data indicate 3 major epidemics of kala-azar in Assam between 1875 and 1950, with inter-epidemic periods of 30-45 and 20 years. This deviates from the popular view of regular cycles with a 10-20 year period. A deterministic mathematical model of kala-azar is used to find the simplest explanation for the timing of the 3 epidemics, paying particular attention to the role of extrinsic (drugs, natural disasters, other infectious diseases) versus intrinsic (host and vector dynamics, birth and death rates, immunity) processes in provoking the second. We conclude that, whilst widespread influenza in 1918-1919 may have magnified the second epidemic, intrinsic population processes provide the simplest explanation for its timing and synchrony throughout Assam. The model also shows that the second inter-epidemic period is expected to be shorter than the first, even in the absence of extrinsic agents, and highlights the importance of a small fraction of patients becoming chronically infectious (with post kala-azar dermal leishmaniasis) after treatment during an epidemic.