Hypoxia increases expression of selected blood–brain barrier transporters GLUT-1, P-gp, SLC7A5 and TFRC, while maintaining barrier integrity, in brain capillary endothelial monolayers

Background Brain capillary endothelial cells (BCECs) experience hypoxic conditions during early brain development. The newly formed capillaries are tight and functional before astrocytes and pericytes join the capillaries and establish the neurovascular unit. Brain endothelial cell phenotype markers P-gp (ABCB1), LAT-1(SLC7A5), GLUT-1(SLC2A1), and TFR(TFRC) have all been described to be hypoxia sensitive. Therefore, we hypothesized that monolayers of BCECs, cultured under hypoxic conditions, would show an increase in LAT-1, GLUT-1 and TFR expression and display tight endothelial barriers. Methods and results Primary bovine BCECs were cultured under normoxic and hypoxic conditions. Chronic hypoxia induced HIF-1α stabilization and translocation to the nucleus, as judged by immunocytochemistry and confocal laser scanning imaging. Endothelial cell morphology, claudin-5 and ZO-1 localization and barrier integrity were unaffected by hypoxia, indicating that the tight junctions in the BBB model were not compromised. SLC7A5, SLC2A1, and TFRC-mRNA levels were increased in hypoxic cultures, while ABCB1 remained unchanged as shown by real-time qPCR. P-gp, TfR and GLUT-1 were found to be significantly increased at protein levels. An increase in uptake of [3H]-glucose was demonstrated, while a non-significant increase in the efflux ratio of the P-gp substrate [3H]-digoxin was observed in hypoxic cells. No changes were observed in functional LAT-1 as judged by uptake studies of [3H]-leucine. Stabilization of HIF-1α under normoxic conditions with desferrioxamine (DFO) mimicked the effects of hypoxia on endothelial cells. Furthermore, low concentrations of DFO caused an increase in transendothelial electrical resistance (TEER), suggesting that a slight activation of the HIF-1α system may actually increase brain endothelial monolayer tightness. Moreover, exposure of confluent monolayers to hypoxia resulted in markedly increase in TEER after 24 and 48 h, which corresponded to a higher transcript level of CLDN5. Conclusions Our findings collectively suggest that hypoxic conditions increase some BBB transporters' expression via HIF-1α stabilization, without compromising monolayer integrity. This may in part explain why brain capillaries show early maturation, in terms of barrier tightness and protein expression, during embryogenesis, and provides a novel methodological tool for optimal brain endothelial culture.

Identification and Characterization of an Early Leaf Senescence Gene ELS1 in Soybean

Early leaf senescence phenotype in soybean could be helpful to shorten the maturation period and prevent green stem disorder. From a high-density mutation library, we identified two early leaf senescence soybean mutant lines, els1-1 (early leaf senescence 1) and els1-2. The chlorophyll contents of both els1-1 and els1-2 were low in pre-senescent leaves. They degraded rapidly in senescent leaves, revealing that ELS1 is involved in chlorophyll biosynthesis during leaf development and chlorophyll degradation during leaf senescence. The causal mutations in els1 were identified by next-generation sequencing-based bulked segregant analysis. ELS1 encodes the ortholog of the Arabidopsis CaaX-like protease BCM1, which is localized in chloroplasts. Soybean ELS1 was highly expressed in green tissue, especially in mature leaves. The accumulation of photosystem I core proteins and light-harvesting proteins in els1 was low even in pre-senescent leaves, and their degradation was accelerated during leaf senescence. These results suggest that soybean ELS1 is involved in both chlorophyll synthesis and degradation, consistent with the findings in Arabidopsis BCM1. The gene els1, characterized by early leaf senescence and subsequent early maturation, does not affect the flowering time. Hence, the early leaf senescence trait regulated by els1 helps shorten the harvesting period because of early maturation characteristics. The els1-1 allele with weakly impaired function of ELS1 has only a small effect on agricultural traits and could contribute to practical breeding.

The effect of bio-banding on academy soccer player passing networks: Implications of relative pitch size

The primary aims of this study were to examine the effects of bio-banding players on passing networks created during 4v4 small-sided games (SSGs), while also examining the interaction of pitch size using passing network analysis compared to a coach-based scoring system of player performance. Using a repeated measures design, 32 players from two English Championship soccer clubs contested mixed maturity and bio-banded SSGs. Each week, a different pitch size was used: Week 1) small (36.1 m2 per player); week 2) medium (72.0 m2 per player); week 3) large (108.8 m2 per player); and week 4) expansive (144.50 m2 per player). All players contested 12 maturity (mis)matched and 12 mixed maturity SSGs. Technical-tactical outcome measures were collected automatically using a foot-mounted device containing an inertial measurement unit (IMU) and the Game Technical Scoring Chart (GTSC) was used to subjectively quantify the technical performance of players. Passing data collected from the IMUs were used to construct passing networks. Mixed effect models were used with statistical inferences made using generalized likelihood ratio tests, accompanied by Cohen’s local f2 to quantify the effect magnitude of each independent variable (game type, pitch size and maturation). Consistent trends were identified with mean values for all passing network and coach-based scoring metrics indicating better performance and more effective collective behaviours for early compared with late maturation players. Network metrics established differences (f2 = 0.00 to 0.05) primarily for early maturation players indicating that they became more integral to passing and team dynamics when playing in a mixed-maturation team. However, coach-based scoring was unable to identify differences across bio-banding game types (f2 = 0.00 to 0.02). Pitch size had the largest effect on metrics captured at the team level (f2 = 0.24 to 0.27) with smaller pitch areas leading to increased technical actions. The results of this study suggest that the use of passing networks may provide additional insight into the effects of interventions such as bio-banding and that the number of early-maturing players should be considered when using mixed-maturity playing formats to help to minimize late-maturing players over-relying on their early-maturing counterparts during match-play.

Identification of a Wheat-Psathyrostachys huashanica 7Ns Ditelosomic Addition Line Conferring Early Maturation by Cytological Analysis and Newly Developed Molecular and FISH Markers

Early maturation is an important objective in wheat breeding programs that could facilitate multiple-cropping systems, decrease disaster- and disease-related losses, ensure stable wheat production, and increase economic benefits. Exploitation of novel germplasm from wild relatives of wheat is an effective means of breeding for early maturity. Psathyrostachys huashanica Keng f. ex P. C. KUO (2n=2x=14, NsNs) is a promising source of useful genes for wheat genetic improvement. In this study, we characterized a novel wheat-P. huashanica line, DT23, derived from distant hybridization between common wheat and P. huashanica. Fluorescence in situ hybridization (FISH) and sequential genomic in situ hybridization (GISH) analyses indicated that DT23 is a stable wheat-P. huashanica ditelosomic addition line. FISH painting and PCR-based landmark unique gene markers analyses further revealed that DT23 is a wheat-P. huashanica 7Ns ditelosomic addition line. Observation of spike differentiation and the growth period revealed that DT23 exhibited earlier maturation than the wheat parents. This is the first report of new earliness per se (Eps) gene(s) probably associated with a group 7 chromosome of P. huashanica. Based on specific locus-amplified fragment sequencing technology, 45 new specific molecular markers and 19 specific FISH probes were developed for the P. huashanica 7Ns chromosome. Marker validation analyses revealed that two specific markers distinguished the Ns genome chromosomes of P. huashanica and the chromosomes of other wheat-related species. These newly developed FISH probes specifically detected Ns genome chromosomes of P. huashanica in the wheat background. The DT23 line will be useful for breeding early maturing wheat. The specific markers and FISH probes developed in this study can be used to detect and trace P. huashanica chromosomes and chromosomal segments carrying elite genes in diverse materials.

Down-regulation of collagen XI during late post-natal corneal development is followed by up-regulation after injury

Collagen XI plays a role in nucleating collagen fibrils and in controlling fibril diameter. The aim of this research is to elucidate the role that collagen XI plays in corneal fibrillogenesis during development and following injury. The temporal and spatial expression of collagen XI was evaluated in C57BL/6 wild type (WT) mice. For wound healing studies in adult mice, stromal injuries were created using techniques that avoid caustic chemicals. The temporal expression and spatial localization of collagen XI was studied following injury in a Col11a1 inducible knockout mouse model. We found that collagen XI expression occurs during early maturation and is upregulated after stromal injury in areas of regeneration and remodeling. Abnormal fibrillogenesis with new fibrils of heterogenous size and shape occurs after injury in a decreased collagen XI matrix. In conclusion, we found that collagen XI is expressed in the stroma during development and following injury in adults. Collagen XI is a regulator of collagen fibrillogenesis in regenerating corneal tissue.

Disease resistance of improved MR220 lines against Pyricularia oryzae Cavara and their preliminary agronomic performance

Blast disease caused by Pyricularia oryzae is one of the most destructive fungal diseases of rice in Malaysia. Utilisation of resistant varieties is the most efficient management approach towards reducing yield losses. The line IRTP21683 with the Pi9 gene has shown strong resistance against the isolate MPO988.3 of pathotype P_0.0, the most prevalent P. oryzae pathotype in Malaysia. Crossing of IRTP21683 was undertaken with the recurrent parent MR220, a susceptible elite Malaysian rice variety, using a marker assisted backcrossing technique with two simple sequence repeat markers, RM19776 and RM7311, as the tag for the Pi9 gene. Twenty BC₃F₄ lines with the Pi9 gene were resistant when challenged with MPO 988.3. The cluster analysis based on seven agronomic parameters showed that the resistant BC₃F₄ lines could be divided into four groups, of which the members in group 1 and 2 have shown comparable or better performance than MR220. Five lines in group 1, B220PI9-3-48, B220PI9-3-76, B220PI9-3-77, B220PI9-3-79 and B220PI9-3-82 showed outstanding yield performance with early maturation.

Source for the breeding of soft spring wheat in the conditions of Kirov region

In the conditions of the Kirov region, 375 samples of soft spring wheat of various ecological and geographical origin (from 30 countries) of the Federal Research Centre N. I. Vavilov All-Russian Institute of Plant Genetic Resources (VIR) collection were studied. The assessment of the source material by yield, duration of the growing season, plant height, lodging resistance, productivity of the ear and plant, 1000-grain mass is presented in the form of a 9-point system of the severity of the trait for three years of field study. As sources for breeding, varieties that combine high yield (5.0...9.0 points) with a high level of economically valuable traits (7.0...9.0 points) were identified. The maximum average point (9.0) for yield capacity was noted by the variety Karabalykskaya 91 (Kazakhstan). The optimal height (53.9...74.0 cm) and lodging resistance were noted in the varieties: Estivum 155 (Russia), Kvorum (Ukraine), Epos, Ethos, Nandu, Schenk and KVS Akvilon (Germany), AC Phil and PT-741 (Canada), Hybrid (Mexico), Leguan (Czech), PS-133 (China) and SSL 25-2 (USA). Early maturation was distinguished in the domestic varieties Skala and Iren (80...87 days). The samples Line 3691h, Izida, FPCh-Ppd-m, Gerakl, Sibirskaya 16 (Russia), Kvorum (Ukraine), Attis and Nandu (Germany), Musket (England) were distinguished by the length (7.3...8.8 cm) and the number of grain of the main ear (32.1...39.9 pcs.). The varieties Voronezhskaya 16, Gerakl, Saratovskaya 72, Saratovskaya 73, Sibirskaya 16, Serebristaya, FPCh-Ppd-m and Ekada 6 (Russia), Anshlag and Kvorum (Ukraine), AC Gabrieland Hoffman (Canada), Attis and Nandu (Germany) were distinguished by a high grain weight per ear (1.28...1.58 g). The varieties Zakamskaya (Russia), Rassvet (Belarus), AC Cadillac (Canada) and PS-95 (China) are recommended as sources of high protein content. It is shown that the wheat yield in the region is closely related to the plant height (r = 0.67), the ear and plant productivity elements (r = 0.24...0.41), the protein content (r = -0.49) and does not depend on the 1000-grain mass (r = 0.04). Sixteen samples were identified with complex resistance to dominating fungal diseases. Samples Altajskaya 110, Kinelskaya 61, Line 2, Lutescens 30 and Estivum V313 (Russia) combined high yield with aluminum resistance. Varieties Altajskaya 100, Baganskaya 95, Line 3691h, Novosibirskaya 20 and Estivum 155 (Russia), Klein Vencedor (Argentina) and NOS Norko (Germany) were characterized by high yield and drought resistance. Based on the obtained results, a bank of sources of economically valuable traits was created, which allows to involve genotypes adapted to the conditions of the region in the breeding process.

Terminal selectors organize postmitotic chromatin accessibility for acquisition of serotonergic identity

Assembly of transcriptomes encoding unique neuronal identities requires selective accessibility of regulatory inputs to cis-regulatory sequences in nucleosome-embedded chromatin. Yet the mechanisms involved in shaping postmitotic neuronal chromatin are poorly understood. Here we used ATAC-seq, ChIPmentation, and single-cell analyses to show that unique distal enhancers and super-enhancers define the Pet1 neuron lineage that generates serotonin (5-HT) neurons. Heterogeneous single cell chromatin landscapes are established early in postmitotic Pet1 neurons and reveal the regulatory programs driving Pet1 neuron subtype identities. Terminal selectors, Pet1 and Lmx1b, control chromatin accessibility in Pet1 neurons to select enhancers for 5-HT neurotransmission and synaptogenesis. In addition, these factors are required to maintain chromatin accessibility during early maturation suggesting that postmitotic open chromatin is unstable and requires continuous terminal selector input. Together our findings reveal a previously unrecognized function of terminal selectors in organizing postmitotic accessible chromatin for the development of specialized neuronal identities.

ON FERTILITY OF BLACK-BACKED SHAD ALOSA KESSLERI KESSLERI (GRIMM, 1887)

The article highlights the results of studying the individual absolute fertility (IAF) of black-backed shad Alosa kessleri kessleri (Grimm, 1887). The data were obtained during the species’ spawning run in the Volga River. A positive correlation was established between IAF and the linear indicators, weight and age of producers. IAF ranged from 38.0 to 333.9 thousand eggs. The average absolute fecundity of herring varied from 38.8 to 259.2 thousand eggs. In the 1970s black-backed shad of 26-42 cm long had fluctuations in the average absolute fecundity from 88.4 to 258.5 thousand eggs. At the present stage, the size range of spawning shoals suggests that maturity in black-backed shad occurs at a body length of 22-45 cm, the reproductive capacity is regulated by early maturation. The results of studies on the fertility of black-backed herring of various lengths in 1 g of ovary have been obtained. There has been registered a wide range of fluctuations in the number of eggs (from 4033 to 14147 pcs). Differences in the size of ovaries indicate a qualitative definiteness of the physiological readiness of females in the pre-spawning period. The fluctuations in the average individual absolute fertility were revealed depending on body weight and age. Fertility in one-size groups and the reproductive capacity of the population as a whole change due to habitat conditions. The conditional indicator of the population fertility was calculated. The results of the obtained fertility data compared with the data of previous years indicate a 1.5-fold decrease in reproductive capacity of the population at the present time, which indicates a stressed state of the reproductive capacity of female species

FSH levels and testicular volumes are associated with the severity of testicular histopathology in men with non-obstructive azoospermia

Purpose The purpose of this study is to assess a potential association between FSH levels and testicular volumes with the severity of testicular histopathology on testicular biopsy in men with non-obstructive azoospermia (NOA) undergoing microdissection testicular sperm extraction (microTESE). Methods A retrospective chart review was performed from the electronic health records of men who underwent microTESE with NOA. Results Eighty-six men with NOA underwent microTESE with concomitant testicular biopsy for permanent section to assess the testicular cellular architecture. The histopathological patterns were categorized by severity indicating the odds of sperm retrieval into 2 categories. The unfavorable category included Sertoli cell only pattern and early maturation arrest (n = 50) and the favorable category included late maturation arrest and hypospermatogenesis patterns (n = 36). In the men with unfavorable histopathologic patterns, the mean FSH level was 22.9 ± 16.6 IU/L, and the mean testicular volume was 10.4 ± 6.0 cc. This was in comparison to men with favorable histopathologic patterns revealing a mean FSH level of FSH 13.3 ± 12.0 with a mean testicular volume of 13.3 ± 5.9 cc. There was a statistically significant higher FSH level in men with unfavorable histopathology than favorable (p = 0.004) as well as a significant smaller mean testicular volume in men with unfavorable histopathology (p = 0.029). Conclusions Higher serum FSH levels and smaller testicular volumes are associated with more severe testicular histopathological patterns in men with NOA.