Lines R1 Research Articles

<i>PD-L1</i> and <i>PD-L2</i> gene expression in human glioblastoma cells resistant to chemo- and radiotherapy

Membrane molecules PD-L1 and PD-L2, ligands of T lymphocytes PD1 receptor, perform immunoregulatory functions. Their binding to the receptor leads to inhibition of proliferation, reduction of cytokine production, cytotoxic response, and apoptosis of T lymphocytes. The cells of many tumors, regardless of their histogenesis, express PD-L1 molecules, thus limiting the development of an anti-tumor immune response. Glioblastomas are highly malignant recurrent tumors of the central nervous system. The main sources of glioblastoma recurrence are resistant tumor cells initially present in gliomas with heterogeneous cellular composition, as well as resistant cells that are formed during therapy. Increasing the dose of cytostatic drugs or radiation during relapse therapy is not effective in glioblastomas. It has been shown for a number of tumors, including ovarian cancer and non-small cell lung cancer, that drugs preventing PD-L1/PD1 interaction are effective in the treatment of neoplasms resistant to chemo- and radiotherapy. Immunotherapy using drugs that inhibit the binding of PD-L molecules to their receptor is considered as a way to overcome the resistance of glioblastomas to therapy. The aim of this work was to assess the level of PD-L1 and PD-L2 gene expression in resistant glioblastoma cells lines A172, R1, T2 and T98G, which resumed proliferation after exposure to the maximum for each line, sublethal doses of cytostatic drugs (fotemustine and temozolomide), fractionated or single gamma irradiation. A172 line belongs to glioblastomas that are highly sensitive to these influences, T98G is a highly resistant cell line, while R1 and T2 lines occupy an intermediate position. In intact glioblastoma A172, R1, and T2 cells the level of PD-L1 and PD-L2 gene expression was equally high, while in T98G cells it was significantly lower. Exposure of A172 and R1 glioblastoma lines to cytostatic drugs or irradiation did not significantly change the level of PD-L1 and PD-L2 genes expression typical for intact cells. In T2 glioblastoma cells, and especially in T98G cells, a significant increase in expression of these genes was found, most pronounced for PD-L2 gene. This increase in expression may indicate an enhanced malignancy of resistant T2 and T98G cells. High expression of the genes responsible for the production of PD-L1 and PD-L2, which limit the cytotoxic response against tumor cells, is a prerequisite for the use of drugs targeted against PD-L1 and PD-L2 for the elimination of resistant cells in glioblastoma.

Diode pumped compact single frequency cw ruby laser

Investigations on 405 nm diode-pumped cw ruby lasers operated in less than 3 mm plane-parallel resonators are reported. With 2.5 mm long ruby crystals TEM00 emission with output powers up to 40 mW are achieved. With an uncoated thin etalon of 0.13 mm thickness, single frequency emission on both ruby lines R1 and R2 is possible. With the piezoelectric shifting of one resonator mirror and corresponding tilting of the etalon, single frequency tuning of up to 400 GHz has been achieved. Details of the laser system are presented, and potential applications will be discussed.

Electroluminescence of Cr3+ and pseudo-Stark effect in β-Ga2O3 Schottky barrier diodes

Bright, red electroluminescence is generated in reverse biased Schottky barrier diodes (SBDs) based on β-Ga2O3 single crystals codoped with chromium and silicon. It is due to intracenter transitions of octahedrally coordinated Cr3+ ions excited by electron impact in the depletion layer of the diodes. The electroluminescence spectrum around 700 nm in wavelength is nearly identical to the spectrum of the well-known photoluminescence of Cr3+ in β-Ga2O3, which is featured by the two lines R1 and R2. In contrast to the photoluminescence, however, in the electroluminescence, each of the R1 and R2 lines is additionally split by 1.5–3 meV. Since the R1/R2 lines correspond to transitions from two Kramers-degenerate states (split 2E excited state) to the ground state 4A2, this splitting cannot be ascribed to the normal Stark effect in the strong electric field of the Schottky barrier diode's depletion layer. Instead, we explain the splitting by the pseudo-Stark effect that occurs because the Cr3+ ions replace Ga3+ at two kinds of energetically equivalent octahedral sites that differ crystallographically by an inversion at the Cr ion. Superposition of the dominating R1 doublet radiation would result in a tunable beat frequency of about 0.4–0.7 THz and might be utilized for a terahertz light source. Moreover, the electroluminescence of chromium is representative of the ability to excite the luminescent states of other transition metals. Hence, high temperature light-emitting SBDs in different colors are a potential application for β-Ga2O3.

The Effect of Fotemustine on Human Glioblastoma Cell Lines

Glioblastomas are a heterogeneous group of tumors of neuroectodermal origin. Tumor cell cultures obtained from patients with glioblastomas differ in morphology and phenotype, as well as genetic characteristics. In the current work we used glioblastoma cell lines A172 and T98G (known since the 1970s) and the new glioblastoma lines R1 and T2 (obtained in 2010 from surgical material of patients). The purpose of this study was to evaluate the effect of the alkylating antitumor drug fotemustine (FM) after single and repeated exposure on the survival and proliferation of these glioblastoma cells, as well as the effect of FM on gene expression of growth factors, extracellular matrix proteins, mesenchymal cell markers, and transcription factor Zeb1. The effect of FM on A172 and R1 glioblastoma cells differs from its effect on T98G and T2 cells. LD50 was in an amount of 60 μg/mL for A172 cells and 250 μg/mL for T98G cells. Single exposure of A172 and R1 cell lines to 100 μg/mL of FM resulted in the death of all cells in culture. A172 and R1 cell exposure to sublethal doses of FM caused decreased activity of all studied genes. After incubation of T98G and T2 glioblastoma cells with FM in sublethal doses we identified two cell populations: proliferating cells and nonproliferating growth-arrested cells. Cell exposure to FM in doses over 300 μg/mL led to the predominance of growth-arrested cells in populations. These cells remained viable for 1.5 months. The effect of FM on T98G and T2 cell lines was accompanied by increased activity of all the studied genes. In conclusion, a population of growth-arrested, drug-resistant cells has been revealed in T98G and T2 cell cultures after exposure to FM. These cells exhibit high expression of genes encoding factors associated with the epithelial–mesenchymal transition. We discuss the possible impact of nonproliferating cells actively producing growth factors on the tumor microenvironment.

Evaluation of Environmental Risk Due to Metro System Construction in Jinan, China.

Jinan is a famous spring city in China. Construction of underground metro system may block groundwater seepage, inducing the depletion risk of springs. This paper presents an assessment of the risk due to metro line construction to groundwater in Jinan City using Analytic Hierarchy Process (AHP) and Geographic International System (GIS). Based on the characteristics of hydrogeology and engineering geology, the assessment model is established from the perspectives of surface index and underground index. The assessment results show that the high and very high risk levels of surface index exceed 98% in the north region; and high and very high risk levels of underground index exceed 56% in urban center and southern region. The assessment result also shows that about 14% of the urban area belongs to very high risk level; regions of high risk are 20% in urban area, 9% in Changqing County and 43% in Pingyin County. In the high risk region, metro lines R1 to R3, which are under construction, and metro lines L1 to L5, which are planned, have very high and high risk. Therefore, risk control measures are proposed to protect the groundwater seepage path to spring.

Cytoskeletal mechanisms in positioning of the second-division spindles and meiotic restitution in tobacco (Nicotiana tabacum L.) microsporogenesis.

Microsporogenesis patterns of the polyploid (2n = 4x = 96) and diploid (2n = 2x = 48) Nicotiana tabacum L. (cv. Havana Petit line SR1) plants have been analyzed and compared. Four types of abnormal positions of the second-division spindles-tripolar, parallel, proximal, and fused-have been observed. Of these abnormalities, only tripolar (2.4%) and parallel (1.4%) spindles are observable in diploid plants. As for polyploids, the increased ploidy is accompanied by an increase in the incidence of tripolar (22.8%) and parallel (8.1%) spindle orientations and emergence of two remaining abnormalities (proximal and fused spindles, 3.3%). As has been shown, the spindle position abnormalities in diploid plants have no effect on the meiotic products, whereas both dyads and triads are detectable among the tetrads in polyploid plants. Analysis of cytoskeletal remodeling has allowed for the insight into the role of interzonal radial microtubule system in spindle positioning during the second division. The reason underlying the change in spindle positioning is disturbed polymerization-depolymerization processes and interdigitation of microtubule plus ends within the interzonal cytoskeleton system in late telophase I-interkinesis and prophase II. As has been demonstrated, fused second-division spindles are formed as a result of fused cytoskeletal structures in prophase-prometaphase II in the case when the nuclei are drawn abnormally close to one another.

Relação entre resistência de linhagens tropicais de milho à podridão de espiga e ao acúmulo de fumonisinas provocados por Fusarium verticillioides

A infecção de grãos de milho por Fusarium verticillioides, agente causal da podridão da espiga, pode resultar na produção de micotoxinas do grupo das fumonisinas. A resistência genética é a forma de controle mais eficiente dessa enfermidade. Assim, o objetivo do trabalho foi buscar fontes de resistência em linhagens de milho tropical à F. verticillioides e à produção de fumonisinas. Seis linhagens tropicais de milho, três, pré-classificadas como resistentes e três, pré-classificadas como suscetíveis à F. verticillioides, foram submetidas à inoculação do patógeno e posteriormente, avaliadas quanto à severidade da podridão de espiga, incidência de grãos sintomáticos e concentração de fumonisinas. Os resultados mostraram que as linhagens R1 e R3 apresentaram alta resistência à infecção do patógeno. No entanto, apenas a R3 foi resistente ao acúmulo de fumonisinas. Dessa forma, sugere-se que a ausência de relação entre intensidade da doença e níveis de fumonisinas seja fator inerente desse patossistema. Assim, não é possível assegurar que grãos assintomáticos quanto à infecção por F. verticillioides, estejam livres de contaminação por fumonisinas.

Combining Ability and Breeding Potential of Rapeseed Elite Lines for Pod Shatter Resistance

Pod shatter resistance of rapeseed is of great importance for modern farming practice. In order to determine the combining ability of elite inbred lines and the breeding potential of rapeseed hybrids in terms of pod shatter resistance, analysis of a 6×6 incomplete diallel cross was conducted at two locations. Results showed that a significant variation existed among breeding lines and their F1 hybrids for pod shatter resistant index (SRI), pod length and width. Pod shatter resistance was significantly positively correlated with pod length. The general combining ability (GCA) effects (GCA=1.58) played a more important role than specific combining ability (SCA) effects (SCA=0.20) for pod shatter trait. The elite lines R1, 1019B and 1055B displayed significant positive GCA effects for pod shatter resistance. Four crosses (1019B×R1, 1015B×R1, 6098B×R1, and 8908B×R1) with high mean performance and positive SCA effects were recommended for developing new hybrids for mechanical harvest in the middle reaches of the Yangtze River.

Ectopic overexpression of peach GDP-d-mannose pyrophosphorylase and GDP-d-mannose-3′,5′-epimerase in transgenic tobacco

We created transgenic tobacco overexpressing peach GDP-d-mannose pyrophosphorylase (PpGMPH) and GDP-d-mannose-3′,5′-epimerase (PpGME), which are involved in the main l-ascorbate (Asc) biosynthetic pathway in plants. Despite an apparent increase in enzymatic activity in the PpGMPH-overexpressing primary transgenic (T0) lines, no significant increase in Asc pool size was observed in leaves. For PpGME-overexpressing T0 lines, Asc pool size was also unchanged, although transcriptional overexpression was confirmed. To reinforce the biosynthetic pathway from d-mannose-1-phosphate to GDP-l-galactose, we further created PpGMPH- and PpGME-double overexpressing crossed transgenic (dOx T1) lines. Transcriptional overexpression of PpGMPH and PpGME transgenes was confirmed, and the Asc content in the dOx T1 lines was slightly elevated in young leaves, albeit not significantly. Western blot analyses revealed protein amounts of GMPH and GME were similar among dOx T1, parental T0 and non-transgenic (SR1) lines in three tissue types tested: leaves, flower buds and immature fruits. Moreover, a similar amount of GMPH and GME proteins were detected in young leaves (YL) and old leaves (OL), although Asc content in YL is roughly double as that in OL. In vitro assay using recombinant PpGME suggested the equilibrium of GME reaction is unfavorable to forward Asc biosynthesis at this step. Finally, to investigate substrate availability in relation to Asc pool size, we added exogenous GDP-d-mannose (5 mM) or l-galactose (5 mM) to tobacco leaf discs; the former resulted in no increase in Asc content, whereas a significant increase was observed in the latter, not only in the dOx T1 lines, but also in the original line SR1. Similar treatments were also conducted under high light, which resulted in a further increase in the increment of Asc content in the l-galactose-fed discs irrespective of transgene expression level. Taken together, both l-galactose availability and light rather than upper biosynthetic gene expression levels of GMPH/GME are critical determinant of Asc content in tobacco leaves, indicating complex modulation of this compound.

An ultrastructural study of microsporogenesis in tobacco line SR1

This article provides an ultrastructural atlas of microsporogenesis in the tobacco model line SR1. The stages of cell-wall remodeling and reorganization of the intercellular channels, accompanying this process, are reported for the microspore mother cells. The meiotic changes in the cell nucleus and cytoplasm are traced. The appearance of single-, double-, or multi-membrane nuclear vacuoles in microspore mother cells and their further elimination from the nucleus are for the first time described for the genus Nicotiana as well as deviations from a normal course for this process. Intercellular chromatin migration (cytomixis) was observed in the microsporogenesis of the line SR1 and behavior of the nuclear vacuoles within the cytomictic nucleus was described for the first time. The enzymatic activity of spherosome-like vesicles in the tobacco microsporogenesis is discussed. The features of microsporogenesis in the tobacco line SR1 are compared with those of other plant species and its association with the transition from a diploid to a haploid phase of the life cycle is discussed.

Estimation of the outcrossing rate for annual Asian wild rice under field conditions

The Asian wild rice, Oryza rufipogon, has partial outcrossing behavior and shows high levels of genetic variation. To estimate an accurate outcrossing rate of annual form of O. rufipogon, two backcross lines (Lines R1 and R2) between Oryza sativa Nipponbare and O. rufipogon W630 were examined under field conditions. A chromosome survey confirmed that these lines had wild chromosomal segments at more than 92% of marker loci. As for the traits of glume, stamen and pistil, Line R1 showed similar floral morphology as that of O. rufipogon W630, whereas Line R2 had larger glumes. In 2005, 2006 and 2008, a total of 22 backcross plants were planted in the middle of wild rice plots. The successive progenies of each plant were examined using microsatellite markers that could clearly detect self-pollination and outcrossing. The outcrossing rates of Line R1 plants ranged from 4.04% to 25.50% with an average of 10.20%. This indicates that cross-pollination of wild rice is a chance event affected by many environmental factors. The outcrossing rates of Line R2 plants also varied, however, no significant difference was observed between the averages of Lines R1 and R2, suggesting that the glume sizes are not critical for outcrossing ability.

Toll-Like Receptor 2 Mediates Proliferation, Survival, NF-κB Translocation, and Cytokine mRNA Expression in LIF-Maintained Mouse Embryonic Stem Cells

Toll-like receptor (TLR) activation is important in immune responses and in differentiation of hematopoietic stem cells. We detected mRNA expression of TLRs 1, 2, 3, 5, and 6, but not TLRs 4, 7, 8, and 9 in murine (m)ESC line E14, and noted high cell surface protein expression of TLR2, but not TLR4, for mESC lines R1, CGR8, and E14. ESC lines were cultured in the presence of leukemia inhibitory factor (LIF). Pam(3)Cys enhanced proliferation and survival of the 3 ESC lines. In contrast, lipopolysaccharide (LPS) decreased proliferation and survival. Pam(3)Cys and LPS effects on proliferation and survival were blocked by antibody to TLR2, suggesting that effects of both Pam(3)Cys and LPS on these mESC lines were likely mediated through TLR2. E14 ESC line expressed MyD88. Pam(3)Cys stimulation of E14 ESCs was associated with induced NF-kappaB translocation, enhanced phosphorylation of IKK-alpha/beta, and enhanced mRNA, but not protein, expression of tumor necrosis factor-alpha, interferon-gamma, and IL-6. TLR2 activation by Pam(3)Cys or inhibition by LPS was not associated with changes in morphology or expression of alkaline phosphatase, Oct4, SSEA1, KLF4, or Sox2, markers of undifferentiated mESCs. Our studies identify TLR2 as present and functional in E14, R1, and CGR8 mESC lines.

Accumulation of overproduced ferritin in the chloroplast provides protection against photoinhibition induced by low temperature in tobacco plants

Wild-type tobacco plants (Nicotiana tabacum L. cv. Petit Havanna line SR1) and plants transformed with full-length alfalfa ferritin cDNA with the chloroplast transit peptide under the control of a Rubisco small subunit gene promoter (C3 and C8) were cold-treated at 0 degrees C with continuous light (250 micromol m(-2)s(-1)). These transgenic plants had higher chlorophyll content and higher F(v)/F(m) chlorophyll-a fluorescence induction parameters than wild-type plants after 2 or 3d of cold treatment in C3 and C8 transgenic plants, respectively. Thermoluminescence studies on the high-temperature bands suggest that these plants suffered less oxidative damage in comparison to the wild-type genotype. The present experiments provide evidence that transgenic tobacco lines overexpressing alfalfa ferritin, which is accumulated in the chloroplasts, may show higher tolerance to various stress factors, generating ROS including low temperature-induced photoinhibition.

Comparison of mammalian and fish cell line cytotoxicity: impact of endpoint and exposure duration

Comparisons of acute toxic concentrations of chemicals to fish in vivo and cytotoxic concentrations to fish cell lines in vitro reveal rather good correlations of the toxic potencies in vitro and in vivo, but a clearly lower sensitivity of the fish cells. To examine whether the low sensitivity is specific for fish cells, cytotoxic potencies of reference chemicals from the Multicenter Evaluation of In Vitro Cytotoxicity program (MEIC) reported for the fish cell lines R1 and RTG-2 were compared with those obtained with the mouse Balb/c 3T3 cell line. Cytotoxic potencies (EC 50 values) for MEIC reference chemicals were determined with exponentially growing Balb/c 3T3 cells using three different test protocols. To assess both endpoints, cell proliferation and cell survival, EC 50 values were measured for the decrease in final cell protein after 24 and 72 h of exposure and for the reduction of cell protein increase during 24 h of exposure. EC 50 values obtained with the fish cell lines R1 and RTG-2 using cell survival as endpoint were taken from the MEIC data base. The comparison of cytotoxic potencies shows that, in general, the fish cell lines and the mammalian cell line are almost equally sensitive towards the cytotoxic action of chemicals. The mammalian cell line assay, however, becomes considerably more sensitive, by factors of 3.4–8.5, than the fish cell line assays, if cell growth instead of cell survival is used as endpoint. It is concluded, that cell proliferation might be a better endpoint than cell survival and that mammalian cell lines might be suited to assess fish acute toxicity.

Vascular stem resistance to black rot in Brassica oleracea

The stem vascular system of several accessions of cultivated Brassica oleracea L. showed resistance to black rot caused by Xanthomonas campestris pv. campestris (Pammel) Dowson. Resistance prevented systemic spread of the pathogen in the vascular system of stem. It was race nonspecific and effective at the early stage of seedling growth. A major dominant gene (Rs; resistant in stem), responsible for expression of compete stem resistance in adult plants (more than six true leaves), was found in a Chinese kale line SR1. At least one more dominant gene with additive interaction was necessary to provide complete resistance in seedlings. In progeny of a cross with a cabbage line PI436606, kale SR1 stem resistance segregated independently of race-specific resistance from a PI436606. It can be considered as an alternative mechanism of plant defense, and a highly valuable feature for crops systemically infected by black rot during propagation in commercial nurseries.Key words: black rot, Xanthomonas campestris pv. campestris, Brassica oleracea, disease resistance, cabbage, kale, broccoli.

Manipulation of in vivo sorbitol production alters boron uptake and transport in tobacco

Recent evidence that some species can retranslocate boron as complexes with sugar alcohols in the phloem suggests a possible mechanism for enhancing boron efficiency. We investigated the relationship between sugar alcohol (sorbitol) content, boron uptake and distribution, and translocation of foliar-applied, isotopically enriched 10B in three lines of tobacco (Nicotiana tabacum) plants differing in sorbitol production. In tobacco line S11, transformed with sorbitol-6-phosphate dehydrogenase, the production of sorbitol was accompanied by an increase in the concentration of boron in plant tissues and an increased uptake of boron compared with either tobacco line A4, transformed with antisense orientation of sorbitol-6-phosphate dehydrogenase, or wild-type tobacco (line SR1, zero-sorbitol producer). Foliar application of 10B to mature leaves was translocated to the meristematic tissues only in line S11. These results demonstrate that the concentration of the boron-complexing sugar alcohol in the plant tissue has a significant effect on boron uptake and distribution in plants, whereas the translocation of the foliar-applied 10B from the mature leaves to the meristematic tissues verifies that boron is mobile in sorbitol-producing plants (S11) as we reported previously. This suggests that selection or transgenic generation of cultivars with an increased sugar alcohol content can result in increased boron uptake, with no apparent negative effects on short-term growth.

Vascular stem resistance to black rot in <i>Brassica oleracea</i>

The stem vascular system of several accessions of cultivated Brassica oleracea L. showed resistance to black rot caused by Xanthomonas campestris pv. campestris (Pammel) Dowson. Resistance prevented systemic spread of the pathogen in the vascular system of stem. It was race nonspecific and effective at the early stage of seedling growth. A major dominant gene (Rs; resistant in stem), responsible for expression of compete stem resistance in adult plants (more than six true leaves), was found in a Chinese kale line SR1. At least one more dominant gene with additive interaction was necessary to provide complete resistance in seedlings. In progeny of a cross with a cabbage line PI436606, kale SR1 stem resistance segregated independently of race-specific resistance from a PI436606. It can be considered as an alternative mechanism of plant defense, and a highly valuable feature for crops systemically infected by black rot during propagation in commercial nurseries.Key words: black rot, Xanthomonas campestris pv...

Characterization of κ-opioid receptor transcripts expressed by T cells and macrophages

We have found that the immature T cell lines R1.1 and DPK and the macrophage lines P388D 1 and WEHI-3 also express κ-opioid receptor (KOR) mRNA. Characterization of the KOR transcripts in both brain tissue and these T cells has revealed both the normal full-length as well as a truncated form of the mRNA. Our results show that the truncated transcript lacks the second exon. Primary macrophages express this truncated form of the transcript in the absence of detectable levels of the full-length form. These results suggest a degree of heterogeneity in the expression of the opioid receptors which has not previously been reported.

Mapping of a Shattering Resistance Gene in a Mutant Line SR- 5 Induced from an indica Rice Variety, Nan-jing11.

A shattering resistance gene in a mutant line, SR-5, induced from an indica rice variety, Nan-jing11 was mapped on chromosome 3 based on QTL (Quantitative Trait Locus) analysis using RFLP markers, and it was designated as sh4. Seventy eight plants of the F2 population derived from a cross between a javanica (tropical japonica) rice variety Dinalaga with shattering habit and SR-5 were investigated for the degree of shattering and marker genotypes by RFLP. The results of QTL analysis suggested that sh4 was located at the same position as that of an RFLP marker R250 on chromosome 3. In the F2 population, the shattering degree showed a continuous distribution including transgressive segregation covering wider ranges of prone and resistance to shattering compared with parental variations. The variance explained (R2) at sh4 showed a lower value than the heritability (h2) of tbe shattering degree in the F2 population. Based on the results of QTL analysis in the region on chromosome 1 where the shattering gene, sh2 and a single recessive shattering resistance gene in a mutant line SR-1, induced from an indica rice variety Nan-jing11, were localted, it was confirmed that these loci were not related to the segregation in the F2 population. It was also suggested tbat in addition to sh4, other factors may control the trait.

THE PRODUCTION AND TECHNOLOGICAL CHARACTERISTICS OF SOME LINES OF PEPPER VARIETY KURTOVSKA KAPIJA IN THE REGION OF STRUMICA

With the long term production of pepper variety Kurtovska kapija, some biological degeneration has appeared. In the Agricultural Institute of Strumica, work has continued on the individual breeding of 798 lines of pepper from several regions of the R. of Macedonia. Lines S1, S2 and S3 from Strumica and lines R1, R2, R3…..R9 form the Radovis region were taken. The vegetative period lasted over 141 days. The larges values concerning fruit length, compared with the standard, were found with the lines R1 and R2 (13.8 cm). The largest values in the fruit width and weight, compared with the standard, were found with line R9 (5.5 cm/74.2 g). The yield obtained from line R9 was the largest (39.149 kg/ha). Knowing the importance of pepper Kurtovska kapija in the vegetable production, and knowing the needs of the food processing industry in the R. of Macedonia, we can say that the lines of these peppers will soon find a place in our vegetable production.