Involvement of homocastasterone, salicylic and abscisic acids in the regulation of drought and freezing tolerance in doubled haploid lines of winter barley

Abstract Ten doubled haploid (DH) lines of winter barley with an increased range of freezing/drought tolerance were used to identify phytohormones involved in plant stress acclimation. Cold hardening and drought stress were applied at the most critical stages of plant development on young seedlings and heading plants, respectively. The level of the phytohormones was significantly higher at heading, more than 5-fold in respect of salicylic acid (SA) and total brassinosteroids (BRs) and 1.7-fold in the case of abscisic acid (ABA). Moreover, the spectrum of detectable BRs increased from one—homocastasterone (HCS)—found in seedlings to four BRs identified in heading plants [HCS, castasterone (CS), teasterone and dolicholide], with the last one detected for the first time in cereal species. To some extent freezing tolerance seems to be determined by native hormonal status as control seedlings of tolerant DH lines contained 1.4- and 2.3-fold lower amount of ABA and HCS and 2.3-fold higher amount of SA in comparison to freezing-sensitive ones. Such dependency was not observed in heading plants as significant variation in CS content was the only detected difference. Under stress treatments, tolerant DH lines accumulated significantly lower (75–81%) amount of ABA, which probably reflected lower stress intensity resulting from another defence strategy. In contrast, stress-induced significant almost 2-fold increase in HCS/CS and 2–3-fold decrease in SA content specific for tolerant DH lines of barley suggest the involvement of these molecules in freezing/drought defence. Detected correlations suggest their interaction with nonspecific peroxidase and low molecular weight antioxidants.

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Screening of camelina (Camelina sativa L.) doubled haploid lines for freezing tolerance in the seedling stage

Genetika ◽

10.2298/gensr1701173s ◽

2017 ◽

Vol 49 (1) ◽

pp. 173-181 ◽

Cited By ~ 6

Author(s):

Jahad Soorni ◽

Seyed Kazemitabar ◽

Danial Kahrizi ◽

Ali Dehestani ◽

Nadali Bagheri

Keyword(s):

Freezing Tolerance ◽

Doubled Haploid ◽

Large Scale ◽

Principal Component ◽

Camelina Sativa ◽

Stress Conditions ◽

Freezing Stress ◽

Doubled Haploid Lines ◽

Survival Percentage ◽

Dh Lines

Freezing stress is an important abiotic stress that limiting the yield and the spatial distribution of many important crops. This study was undertaken to screen 136 doubled haploid (DH) lines of camelina (Camelina sativa L.) along with four canola (Brassica napus) cultivars (Hyola 401, Lord, Roska and Cascade) as experimental control under freezing stress conditions (-14?C for 6 h) to identify lines with high or low level of tolerance to freezing stress for further studies. First, a protocol was developed for large scale screening of camelina germplasm under freezing stress conditions. For this purpose, an experiment with different freezing temperatures (-5, -10, -15 and -20?C) was conducted to find an appropriate temperature that discriminated best between genotypes (i.e. the LT50 temperature). The LT50 values for camelina lines were varied between -10.2 and -17.1?C with an average of -13.94?C for all of the camelina lines. Therefore, we selected the -14?C exposure for 6 h as an appropriate temperature to screening of camelina lines. The principal components of measured parameters (LT50, survival percentage, relative conductivity and scoring) was using principal component analysis that determine freezing-tolerant and freezing-sensitive lines. Among 136 doubled haploid lines, some lines (58, 62 and 101) had higher level of freezing tolerance and some of them (8, 16, 32, 91 and 107) were freezing sensitive. The selected lines in a preliminary freezing screening are useful for further evaluations.

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Mating of 2 Laboratory Saccharomyces cerevisiae Strains Resulted in Enhanced Production of 2-Phenylethanol by Biotransformation of L-Phenylalanine

Journal of Molecular Microbiology and Biotechnology ◽

10.1159/000455169 ◽

2017 ◽

Vol 27 (2) ◽

pp. 81-90 ◽

Cited By ~ 12

Author(s):

Jolanta Mierzejewska ◽

Aleksandra Tymoszewska ◽

Karolina Chreptowicz ◽

Kamil Krol

Keyword(s):

Saccharomyces Cerevisiae ◽

Batch Culture ◽

Fold Increase ◽

Biotechnological Production ◽

Aromatic Alcohol ◽

Enhanced Production ◽

Yeast Strains ◽

First Time

2-Phenylethanol (2-PE) is an aromatic alcohol with a rosy scent which is widely used in the food, fragrance, and cosmetic industries. Promising sources of natural 2-PE are microorganisms, especially yeasts, which can produce 2-PE by biosynthesis and biotransformation. Thus, the first challenging goal in the development of biotechnological production of 2-PE is searching for highly productive yeast strains. In the present work, 5 laboratory Saccharomyces cerevisiae strains were tested for the production of 2-PE. Thereafter, 2 of them were hybridized by a mating procedure and, as a result, a new diploid, S. cerevisiae AM1-d, was selected. Within the 72-h batch culture in a medium containing 5 g/L of <smlcap>L</smlcap>-phenylalanine, AM1-d produced 3.83 g/L of 2-PE in a shaking flask. In this way, we managed to select the diploid S. cerevisiae AM1-d strain, showing a 3- and 5-fold increase in 2-PE production in comparison to parental strains. Remarkably, the enhanced production of 2-PE by the hybrid of 2 yeast laboratory strains is demonstrated here for the first time.

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Production and nutritional value of the wheat silage managed with different cutting systems

Semina Ciências Agrárias ◽

10.5433/1679-0359.2017v38n1p335 ◽

2017 ◽

Vol 38 (1) ◽

pp. 335 ◽

Cited By ~ 1

Author(s):

Rodolfo Carletto ◽

Mikael Neumann ◽

Danúbia Nogueira Figueira ◽

Guilherme Fernando Mattos Leão ◽

Egon Henrique Horst ◽

...

Keyword(s):

Mass Production ◽

Block Design ◽

Neutral Detergent Fiber ◽

Control Treatment ◽

Lower Amount ◽

Mineral Matter ◽

Winter Cereal ◽

Winter Cereals ◽

Cereal Species ◽

Significant Difference

Winter cereals are mainly used for human and animal nutrition and several studies are now exploring their potential as conserved forage (hay and silage). Among the winter cereal species available for cultivation in the south of Brazil, which is major winter cereals producer in the country, the wheat cv. BRS Umbu has attracted special attention. However, few studies have investigated the potential of this cultivar for silage production. The aim of this study was to evaluate the production and quality of silage from the dual-purpose wheat, BRS Umbu, subjected to different cut managements treatments: T1 - without cut (control), T2 - one cut and T3 - two cuts. Each plot represented an experimental unit in a randomized block design, with 5 replicates per treatment. Two representative samples were collected from each plot to determine morphological segmentation (stem, leaf and ear) and dry matter (DM) content of the whole plant and its morphological components. At the time of opening of the silos, food chemical analysis and pH determination were performed. Forage mass production decreased by 26.88% and 67.82%, respectively, with one and two cuts, compared to control. The DM content of the ensiled plant was 49.9 g kg-1 for the control, 54.7 g kg-1 with one cut and 63.2 g kg-1 with two cuts, at the time of ensiling. Management cuts changed the morphological components of the plants, with a lower proportion of stem (28%) in plants subjected to two cuts. The control treatment showed fewer leaves in ensiled plant (9.6%) and intermediate amount of stalk (52.8%), and was significantly different (p < 0.05) to treatment of a cut. Regarding feed chemistry evaluations, no significant difference (p > 0.05) was observed for mineral matter (MM), crude protein (CP) and hemicellulose (HEM) between the different cutting regimens. However, neutral detergent fiber (NDF) and acid detergent fiber (ADF) decreased (p < 0.05) as the number of cuts increased. The control treatment showed higher NDF and ADF content (563.2 and 357.9 g kg-1 DM, respectively) and lower amount of total digestible nutrients (TDN) and net energy of lactation (NEL). However, the estimated milk production (EMP) was superior for this treatment (22,447 l ha-1), demonstrating the high impact of reduced forage mass production with increased number of cuts.

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In vitro and in vivo screening for cold tolerance in winter × spring wheat-derived doubled haploids following the wheat × maize system

Acta Agronomica Hungarica ◽

10.1556/aagr.55.2007.3.2 ◽

2007 ◽

Vol 55 (3) ◽

pp. 273-282

Author(s):

S. Sharma ◽

H. Chaudhary

Keyword(s):

Grain Yield ◽

Cold Tolerance ◽

Spring Wheat ◽

Doubled Haploid ◽

Tetrazolium Chloride ◽

Cold Tolerant ◽

Dh Lines ◽

Ttc Test

Seventy-eight doubled haploid (DH) lines, derived from 21 elite and diverse winter × spring wheat F 1 hybrids, following the wheat × maize system, were screened along with the parental genotypes under in vitro and in vivo conditions for cold tolerance. Under in vitro conditions, the 2,3,5-triphenyl tetrazolium chloride (TTC) test was used to characterize the genotypes for cold tolerance. Based on the TTC test, only one doubled haploid, DH 69, was characterized as cold-tolerant, seven DH and five winter wheat parents were moderately tolerant, while the rest were susceptible. Analysis of variance under in vivo conditions also indicated the presence of sufficient genetic variability among the genotypes (DH lines + parents) for all the yield-contributing traits under study. The correlation and path analysis studies underlined the importance of indirect selection for tillers per plant, harvest index and grains per spike in order to improve grain yield. It was also concluded that selection should not be practised for grain weight per spike as it would adversely affect the grain yield per plant. When comparing the field performance of the genotypes with the in vitro screening parameters, it was concluded that in addition to the TTC test, comprising a single parameter, other physiological and biochemical in vitro parameters should be identified, which clearly distinguish between cold-tolerant and susceptible genotypes and also correlate well with their performance under field conditions.

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Dynamics of humoral immune response in pregnant mares and foals vaccinated with Theileria equi recombinant EMA-2

Pesquisa Veterinária Brasileira ◽

10.1590/1678-5150-pvb-5321 ◽

2018 ◽

Vol 38 (6) ◽

pp. 1105-1109

Author(s):

Alice C. Santos ◽

Fábio P.L. Leite ◽

Ana M. Vianna ◽

Guilherme B. Weege ◽

Ilusca S. Finger ◽

...

Keyword(s):

Immune Response ◽

Humoral Immune Response ◽

Fold Increase ◽

Serum Levels ◽

Vaccination Schedule ◽

Theileria Equi ◽

Humoral Immune ◽

Baseline Values ◽

Antibody Levels ◽

First Time

ABSTRACT: Theileria equi is an infectious hemoprotozoan agent of equine piroplasmosis, a disease that has severe economic and sanitary impact internationally. In addition to its common clinical features, piroplasmosis can cause gestational losses and neonatal damage, which makes neonates susceptible to this disease. The aim of this study was to evaluate the dynamics of humoral immune response to recombinant EMA-2 of T. equi in pregnant mares and foals, as well as the transfer of vaccine antibodies through the colostrum ingested by sucking foals. For vaccine production, the EMA-2 expression gene was cloned and expressed in the yeast species, Pichia pastoris. Thirty-six horses were used, of which 18 were pregnant mares and 18 were foals. The mares were divided into control and vaccinated groups, and the vaccinated group received three doses of rEMA-2 every 21 days starting at 300 days of gestation. Foals from vaccinated and control groups were evaluated until the sixth month of life. The production of antibodies by foals on the rEMA-2 vaccination schedule was also evaluated from the second month of life. Foals in the vaccinated group had received three doses of the vaccine every 21 days. The method used to evaluate serum and colostrum samples was indirect ELISA, and plates were sensitized with the rEMA-2 protein. At the end of the vaccination schedule, vaccinated mares showed a 2.3-fold increase in antibody levels when compared to baseline values. The colostrum of vaccinated mares presented antibody levels of 1.0432±0.33. Foals delivered by vaccinated mares presented levels of antibodies greater than those of foals delivered by control mares after their first time sucking (at about twelve hours after birth). Foals vaccinated in the second month of life showed an 8.3-fold increase in antibody levels when compared to baseline values. The vaccination schedule with rEMA-2 was able to stimulate humoral immunity in pregnant mares. Vaccine immunoglobins were concentrated in the colostrum of vaccinated mares and foals delivered by these mares showed an increase in serum levels of vaccine antibodies after the first-time sucking.

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An improved method for artificial hybridization in annual Medicago species

Australian Journal of Agricultural Research ◽

10.1071/ar9941329 ◽

1994 ◽

Vol 45 (7) ◽

pp. 1329 ◽

Cited By ~ 10

Author(s):

W Pathipanawat ◽

RAC Jones ◽

K Sivasithamparam

Keyword(s):

Success Rate ◽

Fold Increase ◽

Modified Technique ◽

Established Method ◽

Artificial Hybridization ◽

Annual Medicago ◽

Improved Technique ◽

Medicago Species ◽

First Time ◽

Selection Of

An improved technique for successful artificial hybridization in annual medic (Medicago spp.) is described. Using a previously reported method, only four out of seven species were successfully crossed, with the percentage of success ranging from 3 to 22%. Initial modifications to this technique gave a 7-8 fold increase in the successful crossing rate in M. murex and M. polymorpha medic, from 9 to 64% with M. murex and from 10 to 82% with M. polymorpha. Further modifications to the technique resulted in a success rate of 100% in both species. The numbers of seeds per pod obtained from crosses in both species were also increased by using the modified techniques compared to the established method. Selection of larger, more mature flowers, differences in flower cutting position, as well as post pollination position were the main modifications which accounted for the greatly improved success rate. The modified technique was subsequently applied successfully to obtain for the first time inter-specific crosses involving M. polymorphax M. murex, M. polymorphax M. sphaerocarpos, M.murexx M. sphaerocarpos, M. solerolii x M. littoralis/M.truncatula hybrid, M. solerolii x M, tornata, and M. littoralis/M.truncatula hybrid x M. sphaerocarpos.

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Genetics of late maturity α-amylase in a doubled haploid wheat population

Crop and Pasture Science ◽

10.1071/cp09239 ◽

2010 ◽

Vol 61 (2) ◽

pp. 153 ◽

Cited By ~ 14

Author(s):

M. K. Tan ◽

A. P. Verbyla ◽

B. R. Cullis ◽

P. Martin ◽

A. W. Milgate ◽

...

Keyword(s):

Doubled Haploid ◽

Value Added ◽

Wheat Breeding ◽

Wheat Varieties ◽

Breeding Lines ◽

Positive Effects ◽

Wheat Population ◽

B1 Gene ◽

Late Maturity ◽

First Time

Late maturity α-amylase (LMA) in wheat is a defect where high-isoelectric point (pI) α-amylase accumulates in the ripening grain. Wheat genotypes vary in expression from zero to high levels of α-amylase, the latter with detrimental consequences on their use for value-added end products. Expression in each genotype is characterised by varying numbers of grains affected and different levels in each grain. Analysis of a doubled haploid (DH) population (188 lines) from WW1842 × Whistler has identified significant QTL on chromosomes 2DL, 3A, 3B, 3D, 4B, 4D, 5DS and 5BL. The 4B LMA allele (P < 0.0001) from Whistler is closely linked to the QTL for the ‘tall’ allele (P < 0.0001) of the Rht-B1 gene. The 4D LMA QTL (P < 0.0001) in WW1842 co-locates with the QTL for the ‘tall’ allele (P < 0.0001) of the Rht-D1 gene. This study has shown for the first time that a DH cross between two semi-dwarf cultivars with low or no LMA produces ~25% of progeny lines of the ‘tall’ genotypes with a high frequency of LMA. This is attributed to the large additive positive effects from the combination of one recessive ‘tall’ Rht-B1 gene and one recessive ‘tall’ Rht-D1 gene. High-yielding semi-dwarf genotypes with different combinations of Rht-B1 and Rht-D1 alleles which have very low or non-existent LMA expression (e.g. WW1842 and Whistler) may meet industry criteria for registration as commercial wheat varieties. However, when they are used as breeding lines, the cross produces some progeny genotypes with severe levels of LMA. These LMA genotypes comprise the gibberellic acid-sensitive ‘tall’ progenies and a very small proportion of semi-dwarfs. Thus, it is of paramount importance to screen the defect in wheat breeding programs. The suite of QTL identified for LMA will enable the use of marker assisted selection in the pyramiding of the beneficial QTL to maximise yield and minimise (or eliminate) LMA in semi-dwarf genotypes.

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Enhanced Catalytic Dye Decolorization by Microencapsulation of Laccase from P. Sanguineus CS43 in Natural and Synthetic Polymers

Polymers ◽

10.3390/polym12061353 ◽

2020 ◽

Vol 12 (6) ◽

pp. 1353

Author(s):

Natalia Lopez-Barbosa ◽

Ana Lucía Campaña ◽

Juan C. Cruz ◽

Nancy Ornelas-Soto ◽

Johann F. Osma

Keyword(s):

Catalytic Activity ◽

Confocal Microscopy ◽

Fluorescent Dyes ◽

Fold Increase ◽

Dye Decolorization ◽

Reaction Rates ◽

Lower Amount ◽

Poly Vinyl Alcohol ◽

Initial Reaction ◽

Pycnoporus Sanguineus

Polymeric microcapsules with the fungal laccase from Pycnoporus sanguineus CS43 may represent an attractive avenue for the removal or degradation of dyes from wastewaters. Microcapsules of alginate/chitosan (9.23 ± 0.12 µm) and poly(styrenesulfonate) (PSS) (9.25 ± 0.35 µm) were synthesized and subsequently tested for catalytic activity in the decolorization of the diazo dye Congo Red. Successful encapsulation into the materials was verified via confocal microscopy of labeled enzyme molecules. Laccase activity was measured as a function of time and the initial reaction rates were recovered for each preparation, showing up to sevenfold increase with respect to free laccase. The ability of substrates to diffuse through the pores of the microcapsules was evaluated with the aid of fluorescent dyes and confocal microscopy. pH and thermal stability were also measured for encapsulates, showing catalytic activity for pH values as low as 4 and temperatures of about 80 °C. Scanning electron microscope (SEM) analyses demonstrated the ability of PSS capsules to avoid accumulation of byproducts and, therefore, superior catalytic performance. This was corroborated by the direct observation of substrates diffusing in and out of the materials. Compared with our PSS preparation, alginate/chitosan microcapsules studied by others degrade 2.6 times more dye, albeit with a 135-fold increase in units of enzyme per mg of dye. Similarly, poly(vinyl) alcohol microcapsules from degrade 1.7 times more dye, despite an eightfold increase in units of enzyme per mg of dye. This could be potentially beneficial from the economic viewpoint as a significantly lower amount of enzyme might be needed for the same decolorization level achieved with similar encapsulated systems.

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General and specific combining ability of in vitro doubled haploid maize lines in the field

Acta Agronomica Hungarica ◽

10.1556/aagr.58.2010.2.8 ◽

2010 ◽

Vol 58 (2) ◽

pp. 167-177

Author(s):

T. Spitkó ◽

L. SÁgi ◽

J. Pintér ◽

C. Marton ◽

B. Barnabás

Keyword(s):

Doubled Haploid ◽

Combining Ability ◽

Field Performance ◽

Control Line ◽

Regeneration System ◽

Sample Number ◽

Maize Hybrids ◽

Grain Moisture ◽

Dh Lines

The breeding of hybrid maize now has a history of over 100 years. In 1908, George H. Shull was the first to report on the high yields, great uniformity and homogeneity of hybrids derived from a cross between two inbred lines. Following this discovery, consistent self-fertilisation over a period of six to eight generations was found to be an extremely efficient method for developing maize lines. From the mid-1970s, however, with the elaboration of the monoploid ( in vivo ) and microspore culture ( in vitro ) techniques, it became possible to develop homozygous lines within a year.With the help of an efficient plant regeneration system based on anther culture, large numbers of doubled haploid (DH) lines can be produced. In the course of the experiments the seed of DH plants selected over several years was multiplied and crossed with Martonvásár testers, after which the hybrids were included in field performance trials in three consecutive years (2005–2007). The aim was to determine whether the field performance of hybrids developed in this way equalled the mean yield of standards with commercial value. The data also made it possible to calculate the general (GCA) and specific (SCA) combining ability of the parental lines, indicating the usefulness of the parental components in hybrid combinations and expressing the extent to which a given line contributes to yield surpluses in its progeny.A total of 52 maize hybrids were evaluated in the experiments in terms of yield and grain moisture content at harvest. The combinations, resulting from crosses between 12 DH lines, one control line developed by conventional inbreeding and four testers, were found to include hybrids capable of equalling the performance of the standards, and four DH lines were identified as improving the yield level of their progeny. As the experiment was carried out on a very small number of genotypes, the results are extremely promising and suggest that, if the range of genotypes used to develop DH lines is broadened and the sample number is increased, it will be possible in the future to find maize hybrids, developed with in vitro DH parental components, that surpass the performance of commercial hybrids.

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Effects of Selectin Antagonist GMI-1070 on the Activation State of Leukocytes In Sickle Cell Patients Not In Crisis

Blood ◽

10.1182/blood.v116.21.2672.2672 ◽

2010 ◽

Vol 116 (21) ◽

pp. 2672-2672

Author(s):

Scott I Simon ◽

Shannon Chase ◽

Sandra K Larkin ◽

Frans Kuypers ◽

Lori Styles ◽

...

Keyword(s):

Steady State ◽

Sickle Cell ◽

Fold Increase ◽

Employment Equity ◽

Blood Samples ◽

High Affinity ◽

Equity Ownership ◽

First Time ◽

Time Point

Abstract Abstract 2672 It is hypothesized that activated leukocytes play key roles in sickle cell vaso-occlusion by adhering to inflamed venules and capturing circulating platelets and sickle red blood cells. GMI-1070 is a small molecule selectin antagonist which was recently reported to reverse acute vascular occlusion in a humanized sickle cell disease (SCD) mouse model (Chang et al, Blood 2010) presumably by inhibiting E-selectin and its effects on downstream signaling of leukocyte activation. Sickle cell patients express elevated levels of soluble E-selectin (Kato et al, Brit J Haem 2005) activated polymorphonuclear neutrophils (PMN) (Lum et al Amer J Hem 2004) and platelet/monocyte aggregates (PMA) (Wun et al Clin Lab Haem 2002). In this study, the activation state of leukocytes from whole blood samples of sickle cell patients not in crisis before and after infusion of GMI-1070 was evaluated ex vivo. Isolated PMN from normal, healthy volunteers were strongly activated by binding soluble E-selectin/hIg in vitro as determined by a 7-fold increase of the integrin MAC1 (CD11b) and an 8-fold increase in expression of the high affinity form of CD18 detected by antibody 327C. Addition of GMI-1070 completely blocked upregulation of MAC1 and 327C at 50μg/ml and showed pronounced inhibition (79% MAC1; 75% 327C) at 10μg/ml. These in vitro concentrations are consistent with blood levels of GMI-1070 found in sickle cell patients 4 and 8 hours after dosing. A phase 1/2 study was conducted on 10 adult subjects with SCD at steady state. GMI-1070 was given IV at 20mg/kg as a loading dose and at 10 hours a final dose of 10mg/kg was given. Blood samples were drawn from these adults pre-infusion and at 8, 24, and 48 hours after the initial infusion. In some subjects, a blood sample was also drawn at 4 hours post infusion. Activation of PMN's in whole blood samples from subjects was assessed by upregulation of MAC-1, expression of the high affinity CD18 and the loss of CD62L due to shedding of L-selectin determined by flow cytometric analysis of cell surface labeling with fluorescently conjugated antibodies. Of 4 subjects tested, 3 showed increased surface expression of L-selectin, 3 showed decreased expression of MAC-1, and 2 showed decreased expression of high affinity CD11b at the first time point tested (4 or 8hr) after dosing with GMI-1070 suggesting an inhibition of PMN activation in these patients. A functional consequence of monocyte activation is the formation of platelet/monocyte aggregates due to expression of high affinity integrins. Platelet-monocytes aggregates (PMA) in blood were detected using anti-CD11c for monocytes and anti-CD41a for platelets. Treatment of samples with lipopolysaccharide (LPS) was used for positive controls. Intracellular IL-1β was used as a marker of activated monocytes. In 5 patients out of 6 tested with this assay, PMA in the subject's blood were decreased at the first time point after dosing (8hr). These results are consistent with an effect of GMI-1070 on inhibition of activation given its IC50 value for E-selectin (4.3μM), the blood concentration in subjects after dosing, and the serum half life (7.7hr) in steady state sickle cell adults. Conclusions: GMI-1070 significantly inhibited E-selectin-mediated activation of PMNs in vitro as determined by expression of the integrin MAC-1 and high affinity CD18 at 10μg/ml. Similar concentrations of GMI-1070 in sickle cell subjects' blood at 4 and 8 hours after dosing also resulted in a lowered activation state of PMNs identified by reduced expression of cell surface integrin molecules as well as the inhibition of shedding of L-selectin in some cases. A more functional measure of leukocyte activation is the aggregation of platelets on monocyte cell surfaces. In 5 of 6 subjects tested, GMI-1070 reduced PMA 8 hours after dosing. Thus, GMI-1070 not only inhibits E-selectin, but also blocks the expression of downstream integrin adhesion molecules that together play crucial roles in vaso-occlusion by promoting the adhesion to platelets and erythrocytes in the formation of occlusions that block blood flow. The effects of GMI-1070 on the activation state of leukocytes via the inhibition of functional adhesion molecules in steady state sickle cell subjects supports the further evaluation of treatment with GMI-1070 during vaso-occlusive episodes. Disclosures: Simon: GlycoMimetics: Research Funding. Chase:GlycoMimetics:.Kuypers:GlycoMimetics Inc.: Research Funding. Styles:GlycoMimetics: Consultancy, clinical trial sponsorship. Wun:GlycoMimetics Inc.: Consultancy, clinical trial sponsorship. Thackray:GlycoMimetics: Employment, Equity Ownership. Magnani:GlycoMimetics: Employment, Equity Ownership. Off Label Use: The drug (GMI-1070) is not approved for any clinical indication.

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