Genome-wide and functional analysis of late embryogenesis abundant (LEA) genes during dormancy and sprouting periods of kernel consumption apricots (P. armeniaca L. × P. sibirica L.).

Late embryogenesis abundant (LEA) proteins play a crucial role in protecting cells from stress, making them potential contributors to abiotic stress tolerance. This study focuses on apricot (P. armeniaca L. × P. sibirica L.), where a comprehensive genome-wide analysis identified 54 LEA genes, categorized into eight subgroups based on phylogenetic relationships. Synteny analysis revealed 14 collinear blocks containing LEA genes between P. armeniaca × P. sibirica and Arabidopsis thaliana, with an additional 9 collinear blocks identified between P. armeniaca × P. sibirica and poplar. Examination of gene structure and conserved motifs indicated that these subgroups exhibit consistent exon-intron patterns and shared motifs. The expansion and duplication of LEA genes in P. armeniaca × P. sibirica were driven by whole-genome duplication (WGD), segmental duplication, and tandem duplication events. Expression analysis, utilizing RNA-seq data and quantitative real-time RT-PCR (qRT-PCR), indicated induction of PasLEA2-20, PasLEA3-2, PasLEA6-1, Pasdehydrin-3, and Pasdehydrin-5 in flower buds during dormancy and sprouting phases. Coexpression network analysis linked LEA genes with 15 cold-resistance genes. Remarkably, during the four developmental stages of flower buds in P. armeniaca × P. sibirica - physiological dormancy, ecological dormancy, sprouting period, and germination stage - the expression patterns of all PasLEAs coexpressed with cold stress-related genes remained consistent. Protein-protein interaction networks, established using Arabidopsis orthologs, emphasized connections between PasLEA proteins and cold resistance pathways. Overexpression of certain LEA genes in yeast and Arabidopsis conferred advantages under cold stress, including increased pod length, reduced bolting time and flowering time, improved survival and seed setting rates, elevated proline accumulation, and enhanced antioxidative enzymatic activities. Furthermore, these overexpressed plants exhibited upregulation of genes related to flower development and cold resistance. The Y1H assay confirmed that PasGBF4 and PasDOF3.5 act as upstream regulatory factors by binding to the promoter region of PasLEA3-2. PasDOF2.4, PasDnaJ2, and PasAP2 were also found to bind to the promoter of Pasdehydrin-3, regulating the expression levels of downstream genes. This comprehensive study explores the evolutionary relationships among PasLEA genes, protein interactions, and functional analyses during various stages of dormancy and sprouting in P. armeniaca × P. sibirica. It offers potential targets for enhancing cold resistance and manipulating flower bud dormancy in this apricot hybrid.

The superior allele LEA12OR in wild rice enhances salt tolerance and yield.

Soil salinity has negative impacts on food security and sustainable agriculture. Ion homeostasis, osmotic adjustment and reactive oxygen species scavenging are the main approaches utilized by rice to resist salt stress. Breeding rice cultivars with high salt tolerance (ST) and yield is a significant challenge due to the lack of elite alleles conferring ST. Here, we report that the elite allele LEA12OR, which encodes a late embryogenesis abundant (LEA) protein from the wild rice Oryza rufipogon Griff., improves osmotic adjustment and increases yield under salt stress. Mechanistically, LEA12OR, as the early regulator of the LEA12OR-OsSAPK10-OsbZIP86-OsNCED3 functional module, maintains the kinase stability of OsSAPK10 under salt stress, thereby conferring ST by promoting abscisic acid biosynthesis and accumulation in rice. The superior allele LEA12OR provides a new avenue for improving ST and yield via the application of LEA12OR in current rice through molecular breeding and genome editing.

Genome-wide identification of the LEA gene family in Panax ginseng: Evidence for the role of PgLEA2-50 in plant abiotic stress response.

Ginseng frequently encounters environmental stress during its growth and development. Late Embryogenesis Abundant (LEA) proteins play a crucial role in combating adversity stress, particularly against abiotic challenges In this study, 107 LEA genes from ginseng, spanning eight subfamilies, were identified, demonstrating significant evolutionary conservation, with the LEA2 subfamily being most prominent. Gene duplication events, primarily segmental duplications, have played a major role in the expansion of the LEA gene family, which has undergone strong purifying selection. PgLEAs were unevenly distributed across 22 chromosomes, with each subfamily featuring unique structural domains and conserved motifs. PgLEAs were expressed in various tissues, exhibiting distinct variations in abundance and tissue specificity. Numerous regulatory cis-elements, related to abiotic stress and hormones, were identified in the promoter region. Additionally, PgLEAs were regulated by a diverse array of abiotic stress-related transcription factors. A total of 35 PgLEAs were differentially expressed following treatments with ABA, GA, and IAA. Twenty-three PgLEAs showed significant but varied responses to drought, extreme temperatures, and salinity stress. The transformation of tobacco with the key gene PgLEA2-50 enhanced osmoregulation and antioxidant levels in transgenic lines, improving their resistance to abiotic stress. This study offers insights into functional gene analysis, focusing on LEA proteins, and establishes a foundational framework for research on ginseng's resilience to abiotic stress.

Genome-wide characterization of LEA gene family reveals a positive role of BnaA.LEA6.a in freezing tolerance in rapeseed (Brassica napus L.).

BACKGROUND: Freezing stress is one of the major abiotic stresses that causes extensive damage to plants. LEA (Late embryogenesis abundant) proteins play a crucial role in plant growth, development, and abiotic stress. However, there is limited research on the function of LEA genes in low-temperature stress in Brassica napus (rapeseed). RESULTS: Total 306 potential LEA genes were identified in B. rapa (79), B. oleracea (79) and B. napus (148) and divided into eight subgroups. LEA genes of the same subgroup had similar gene structures and predicted subcellular locations. Cis-regulatory elements analysis showed that the promoters of BnaLEA genes rich in cis-regulatory elements related to various abiotic stresses. Additionally, RNA-seq and real-time PCR results indicated that the majority of BnaLEA family members were highly expressed in senescent tissues of rapeseed, especially during late stages of seed maturation, and most BnaLEA genes can be induced by salt and osmotic stress. Interestingly, the BnaA.LEA6.a and BnaC.LEA6.a genes were highly expressed across different vegetative and reproductive organs during different development stages, and showed strong responses to salt, osmotic, and cold stress, particularly freezing stress. Further analysis showed that overexpression of BnaA.LEA6.a increased the freezing tolerance in rapeseed, as evidenced by lower relative electrical leakage and higher survival rates compared to the wild-type (WT) under freezing treatment. CONCLUSION: This study is of great significance for understanding the functions of BnaLEA genes in freezing tolerance in rapeseed and offers an ideal candidate gene (BnaA.LEA6.a) for molecular breeding of freezing-tolerant rapeseed cultivars.

Improving vitrification efficiency of human in vitro matured oocytes by the addition of LEA proteins.

STUDY QUESTION: Can the addition of late embryogenesis-abundant (LEA) proteins as a cryoprotective agent during the vitrification cryopreservation of in vitro matured oocytes enhance their developmental potential after fertilization? SUMMARY ANSWER: LEA proteins improve the developmental potential of human in vitro matured oocytes following cryopreservation, mostly by downregulating FOS genes, reducing oxidative stress, and inhibiting the formation of ice crystals. WHAT IS KNOWN ALREADY: Various factors in the vitrification process, including cryoprotectant toxicity, osmotic stress, and ice crystal formation during rewarming, can cause fatal damage to oocytes, thereby affecting the oocytes developmental potential and subsequent clinical outcomes. Recent studies have shown that LEA proteins possess high hydrophilicity and inherent stress tolerance, and can reduce low-temperature damage, although the molecular mechanism it exerts protective effects is still unclear. STUDY DESIGN, SIZE, DURATION: Two LEA proteins extracted and purified by us were added to solutions for vitrification-warming of oocytes at concentrations of 10, 100, and 200 µg/mL, to determine the optimal protective concentration for each protein. Individual oocyte samples were collected for transcriptomic analysis, with each group consisting of three sample replicates. PARTICIPANTS/MATERIALS, SETTING, METHODS: Immature oocytes were collected from patients who were undergoing combined in vitro fertilization (IVF) treatment and who had met the designated inclusion and exclusion criteria. These oocytes underwent in vitro maturation (IVM) culture for experimental research. A fluorescence microscope was used to detect the levels of mitochondrial membrane potential (MMP), reactive oxygen species (ROS), and calcium in the mitochondria of vitrified-warmed human oocytes treated with different concentrations of LEA proteins, and the protective effect of the protein on mitochondrial function was assessed. The levels of intracellular ice recrystallization inhibition (IRI) in human oocytes after vitrification-warming were characterized by the cryomicroscope, to determine the LEA proteins inhibitory effect on recrystallization. By analyzing transcriptome sequencing data to investigate the potential mechanism through which LEA proteins exert their cryoprotective effects. MAIN RESULTS AND THE ROLE OF CHANCE: The secondary structures of AfrLEA2 and AfrLEA3m proteins were shown to consist of a large number of α-helices and the proteins were shown to be highly hydrophilic, in agreement with previous reports. Confocal microscopy results showed that the immunofluorescence of AfrLEA2-FITC and AfrLEA3m-FITC-labeled proteins appeared to be extracellular and did not penetrate the cell membrane compared with the fluorescein isothiocyanate (FITC) control group, indicating that both AfrLEA2 and AfrLEA3m proteins were extracellular. The group treated with 100 µg/mL AfrLEA2 or AfrLEA3m protein had more uniform cytoplasmic particles and fewer vacuoles compared to the 10 and 200 µg/mL groups and were closest to the fresh group. In the 100 µg/mL groups, MMPs were significantly higher while ROS and calcium levels were significantly lower than those in the control group and were closer to the levels observed in fresh oocytes. Meanwhile, 100 µg/mL of AfrLEA2 or AfrLEA3m protein caused smaller ice crystal formation in the IRI assay compared to the control group treated with dimethylsulphoxide (DMSO) and ethylene glycol (EG); thus, the recrystallization inhibition was superior to that with the conventional cryoprotectants DMSO and EG. Further results revealed that the proteins improved the developmental potential of human oocytes following cryopreservation, likely by downregulating FOS genes and reducing oxidative stress. LIMITATIONS, REASONS FOR CAUTION: The in vitro-matured metaphase II (IVM-MII) oocytes used in the study, due to ethical constraints, may not accurately reflect the condition of MII oocytes in general. The AfrLEA2 and AfrLEA3m proteins are recombinant proteins and their synthetic stability needs to be further explored. WIDER IMPLICATIONS OF THE FINDINGS: LEA proteins, as a non-toxic and effective cryoprotectant, can reduce the cryoinjury of oocytes during cryopreservation. It provides a new promising method for cryopreservation of various cell types. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Key Research and Development Program of China (2022YFC2703000) and the National Natural Science Foundation of China (52206064). The authors declare no competing interest. TRIAL REGISTRATION NUMBER: N/A.

Expression of the Arabidopsis redox-related LEA protein, SAG21 is regulated by ERF, NAC and WRKY transcription factors.

SAG21/LEA5 is an unusual late embryogenesis abundant protein in Arabidopsis thaliana, that is primarily mitochondrially located and may be important in regulating translation in both chloroplasts and mitochondria. SAG21 expression is regulated by a plethora of abiotic and biotic stresses and plant growth regulators indicating a complex regulatory network. To identify key transcription factors regulating SAG21 expression, yeast-1-hybrid screens were used to identify transcription factors that bind the 1685 bp upstream of the SAG21 translational start site. Thirty-three transcription factors from nine different families bound to the SAG21 promoter, including members of the ERF, WRKY and NAC families. Key binding sites for both NAC and WRKY transcription factors were tested through site directed mutagenesis indicating the presence of cryptic binding sites for both these transcription factor families. Co-expression in protoplasts confirmed the activation of SAG21 by WRKY63/ABO3, and SAG21 upregulation elicited by oligogalacturonide elicitors was partially dependent on WRKY63, indicating its role in SAG21 pathogen responses. SAG21 upregulation by ethylene was abolished in the erf1 mutant, while wound-induced SAG21 expression was abolished in anac71 mutants, indicating SAG21 expression can be regulated by several distinct transcription factors depending on the stress condition.

Disordered proteins interact with the chemical environment to tune their protective function during drying.

The conformational ensemble and function of intrinsically disordered proteins (IDPs) are sensitive to their solution environment. The inherent malleability of disordered proteins combined with the exposure of their residues accounts for this sensitivity. One context in which IDPs play important roles that is concomitant with massive changes to the intracellular environment is during desiccation (extreme drying). The ability of organisms to survive desiccation has long been linked to the accumulation of high levels of cosolutes such as trehalose or sucrose as well as the enrichment of IDPs, such as late embryogenesis abundant (LEA) proteins or cytoplasmic abundant heat soluble (CAHS) proteins. Despite knowing that IDPs play important roles and are co-enriched alongside endogenous, species-specific cosolutes during desiccation, little is known mechanistically about how IDP-cosolute interactions influence desiccation tolerance. Here, we test the notion that the protective function of desiccation-related IDPs is enhanced through conformational changes induced by endogenous cosolutes. We find that desiccation-related IDPs derived from four different organisms spanning two LEA protein families and the CAHS protein family, synergize best with endogenous cosolutes during drying to promote desiccation protection. Yet the structural parameters of protective IDPs do not correlate with synergy for either CAHS or LEA proteins. We further demonstrate that for CAHS, but not LEA proteins, synergy is related to self-assembly and the formation of a gel. Our results demonstrate that functional synergy between IDPs and endogenous cosolutes is a convergent desiccation protection strategy seen among different IDP families and organisms, yet, the mechanisms underlying this synergy differ between IDP families.

AmDHN4, a winter accumulated SKn-type dehydrin from Ammopiptanthus mongolicus, and regulated by AmWRKY45, enhances the tolerance of Arabidopsis to low temperature and osmotic stress.

Ammopiptanthus mongolicus, a rare temperate evergreen broadleaf shrub, exhibits remarkable tolerance to low temperature and drought stress in winter. Late embryogenesis abundant (LEA) proteins, a kind of hydrophilic protein with a protective function, play significant roles in enhancing plant tolerance to abiotic stress. In this present study, we analyzed the evolution and expression of LEA genes in A. mongolicus, and investigated the function and regulatory mechanism of dehydrin under abiotic stresses. Evolutionary analysis revealed that 14 AmLEA genes underwent tandem duplication events, and 36 AmLEA genes underwent segmental duplication events Notably, an expansion in SKn-type dehydrins was observed. Expression analysis showed that AmDHN4, a SKn-type dehydrin, was up-regulated in winter and under low temperature and osmotic stresses. Functional analysis showcased that the heterologous expression of the AmDHN4 enhanced the tolerance of yeast and tobacco to low temperature stress. Additionally, the overexpression of AmDHN4 significantly improved the tolerance of transgenic Arabidopsis to low temperature, drought, and osmotic stress. Further investigations identified AmWRKY45, a downstream transcription factor in the jasmonic acid signaling pathway, binding to the AmDHN4 promoter and positively regulating its expression. In summary, these findings contribute to a deeper understanding of the functional and regulatory mechanisms of dehydrin.

Heterologous overexpression of Tawzy1-2 gene encoding an SK3 dehydrin enhances multiple abiotic stress tolerance in Escherichia coli and Nicotiania benthamiana.

MAIN CONCLUSION: The nuclear localized TaWZY1-2 helps plants resist abiotic stress by preserving the cell's ability to remove reactive oxygen species and decrease lipid oxidation under such conditions. In light of the unpredictable environmental conditions in which food crops grow, precise strategies must be developed by crops to effectively cope with abiotic stress and minimize damage over their lifespan. A key component in this endeavor is the group II of late embryogenesis abundant (LEA) proteins, known as dehydrins, which play crucial roles in enhancing the tolerance of plants to abiotic stress. Tawzy1-2 is a dehydrin-encoding gene which is constitutively expressed in various tissues of wheat. However, the biological function of TaWZY1-2 is not yet fully understood. In this study, TaWZY1-2 was isolated and identified in the wheat genome, and its functional role in conferring tolerance to abiotic stresses was detected in both prokaryotic and eukaryotic cells. Results showed that TaWZY1-2 is a nuclear localized hydrophilic protein that accumulates in response to multiple stresses. Escherichia coli cells expressing TaWZY1-2 showed enhanced tolerance to multiple stress conditions. Overexpression of TaWZY1-2 in Nicotiania benthamiana improved growth, germination and survival rate of the transgenic plants exposed to four kinds of abiotic stress conditions. Our results show that Tawzy1-2 transgenic plants exhibit improved capability in clearing reactive oxygen species and reducing lipid degradation, thereby enhancing their resistance to abiotic stress. This demonstrates a significant role of TaWZY1-2 in mitigating abiotic stress-induced damage. Consequently, these findings not only establish a basis for future investigation into the functional mechanism of TaWZY1-2 but also contribute to the expansion of functional diversity within the dehydrin protein family. Moreover, they identify potential candidate genes for crop optimization.

Proteome plasticity during Physcomitrium patens spore germination - from the desiccated phase to heterotrophic growth and reconstitution of photoautotrophy.

The establishment of moss spores is considered a milestone in plant evolution. They harbor protein networks underpinning desiccation tolerance and accumulation of storage compounds that can be found already in algae and that are also utilized in seeds and pollen. Furthermore, germinating spores must produce proteins that drive the transition through heterotrophic growth to the autotrophic plant. To get insight into the plasticity of this proteome, we investigated it at five timepoints of moss (Physcomitrium patens) spore germination and in protonemata and gametophores. The comparison to previously published Arabidopsis proteome data of seedling establishment showed that not only the proteomes of spores and seeds are functionally related, but also the proteomes of germinating spores and young seedlings. We observed similarities with regard to desiccation tolerance, lipid droplet proteome composition, control of dormancy, and ß-oxidation and the glyoxylate cycle. However, there were also striking differences. For example, spores lacked any obvious storage proteins. Furthermore, we did not detect homologs to the main triacylglycerol lipase in Arabidopsis seeds, SUGAR DEPENDENT1. Instead, we discovered a triacylglycerol lipase of the oil body lipase family and a lipoxygenase as being the overall most abundant proteins in spores. This finding indicates an alternative pathway for triacylglycerol degradation via oxylipin intermediates in the moss. The comparison of spores to Nicotiana tabacum pollen indicated similarities for example in regards to resistance to desiccation and hypoxia, but the overall developmental pattern did not align as in the case of seedling establishment and spore germination.

Hormonal contraception for female athletes presents special needs and concerns.

PURPOSE: Oral contraceptives (OCs) are commonly used by female athletes raising concerns regarding the possible adverse effects of OCs on physical performance, musculoskeletal injuries, and bone density. We aimed to review all current studies on the physiological effects of OCs in physically active women. MATERIALS AND METHODS: A review of literature in electronic search in PubMed and Google Scholar databases from December 2002 to December 2022 using relevant keywords. The reference lists of the articles found eligible were also reviewed. RESULTS: Out of 344 articles in the initial database, 54 clinical studies were eligible for inclusion in our literature review. OCs are used by about two-thirds of female athletes. Current research suggests that OCs' effects on endurance performance and muscle strength are mostly reassuring. OCs do not seem to have a major negative impact on bone health or sports injuries. In fact, new data suggests that they may even significantly reduce the risk of anterior cruciate ligament (ACL) injury. CONCLUSIONS: OCs can be safely used by young female athletes, who may also benefit from better menstrual cycle control. OCs offer newly realised protection from ACL injuries. The use of OCs must be carefully individualised according to their preferences, expectations, and experience.

Hormonal contraception can be safely prescribed to active women. The International Olympic Committee advocates a new and wider definition for the 'athlete triad', where low energy availability presents a special challenge for these athletes. New data suggests that the use of oral contraceptives may protect against ACL injury.

Research progress on commonly used visual acuity charts for preschool children / 国际眼科杂志(Guoji Yanke Zazhi)

Preschool age(3-6 years old)is a critical period for visual development, and it is crucial to detect and treat visual problems in preschool children as early as possible. Visual acuity charts are important tools for screening visual issues in children. In China, the commonly used charts are the standard logarithmic visual acuity chart and the pediatric optotype chart, while overseas, the Lea, HOTV, and ETDRS visual acuity charts are frequently employed. Numerous studies have reported the measurability, repeatability, and sensitivity of these three charts in diagnosing visual-related problems in children. However, the application of these three charts is relatively limited in China. This article provides a comprehensive review of the design principles, clinical applications, and characteristics of these three visual acuity charts, so as to better understand their applicability and limitations in preschool children, and provide reference for the selection and improvement of vision examination methods in the future.

The Late Embryogenesis Abundant Proteins in Soybean: Identification, Expression Analysis, and the Roles of GmLEA4_19 in Drought Stress.

Late embryogenesis abundant (LEA) proteins play important roles in regulating plant growth and responses to various abiotic stresses. In this research, a genome-wide survey was conducted to recognize the LEA genes in Glycine max. A total of 74 GmLEA was identified and classified into nine subfamilies based on their conserved domains and the phylogenetic analysis. Subcellular localization, the duplication of genes, gene structure, the conserved motif, and the prediction of cis-regulatory elements and tissue expression pattern were then conducted to characterize GmLEAs. The expression profile analysis indicated that the expression of several GmLEAs was a response to drought and salt stress. The co-expression-based gene network analysis suggested that soybean LEA proteins may exert regulatory effects through the metabolic pathways. We further explored GnLEA4_19 function in Arabidopsis and the results suggests that overexpressed GmLEA4_19 in Arabidopsis increased plant height under mild or serious drought stress. Moreover, the overexpressed GmLEA4_19 soybean also showed a drought tolerance phenotype. These results indicated that GmLEA4_19 plays an important role in the tolerance to drought and will contribute to the development of the soybean transgenic with enhanced drought tolerance and better yield. Taken together, this study provided insight for better understanding the biological roles of LEA genes in soybean.

Considerations of Low Carbohydrate Availability (LCA) to Relative Energy Deficiency in Sport (RED-S) in Female Endurance Athletes: A Narrative Review.

The purpose of this narrative review is to identify health and performance consequences associated with LCA in female endurance athletes. The intake of carbohydrates (CHO) before, during, and after exercise has been demonstrated to support sport performance, especially endurance activities which rely extensively on CHO as a fuel source. However, low energy availability (LEA) and low carbohydrate availability (LCA) are common in female athletes. LEA occurs when energy intake is insufficient compared to exercise energy expenditure, and LEA-related conditions (e.g., Female Athlete Triad (Triad) and Relative Energy Deficiency in Sport (RED-S)) are associated with a myriad of health and performance consequences. The RED-S model highlights 10 health consequences and 10 performance consequences related to LEA. The independent effect of LCA on health and performance has been under-researched, despite current CHO intake being commonly insufficient in athletes. It is proposed that LCA may not only contribute to LEA but also have independent health and performance consequences in athletes. Furthermore, this review highlights current recommendations for CHO intake, as well as recent data on LCA prevalence and menstrual cycle considerations. A literature review was conducted on PubMed, Science Direct, and ResearchGate using relevant search terms (i.e., "low carbohydrate/energy availability", "female distance runners"). Twenty-one articles were identified and twelve met the inclusion criteria. The total number of articles included in this review is 12, with 7 studies illustrating that LCA was associated with direct negative health and/or performance implications for endurance-based athletes. Several studies included assessed male athletes only, and no studies included a female-only study design. Overall, the cumulative data show that female athletes remain underrepresented in sports science research and that current CHO intake recommendations and strategies may fail to consider female-specific adaptations and hormone responses, such as monthly fluctuations in estrogen and progesterone throughout the menstrual cycle. Current CHO guidelines for female athletes and exercising women need to be audited and explored further in the literature to support female athlete health and performance.

Analysis of Lower Extremity Amputations from the SerbVasc Registry.

BACKGROUND: Peripheral arterial disease (PAD) and diabetes are the major causes of lower extremity amputations (LEAs) worldwide. Morbidity and mortality in patients with LEAs are high with an associated significant burden on the global health system. The aim of this article is to report the overall morbidity and mortality rates after major and minor LEAs from the Serbian Vascular Registry (SerbVasc), with an analysis of predictive factors that influenced adverse outcomes. MATERIALS AND METHODS: SerbVasc was created in 2019 as a part of the Vascunet collaboration that is aiming to include all vascular procedures from 21 hospitals in Serbia. Prevalence of diabetes among patients with LEAs, previous revascularization procedures, the degree and the type of foot infection and tissue loss, and overall morbidity and mortality rates were analyzed, with a special reference to mortality predictors. RESULTS: In the period from January 2020 to December 2022, data on 702 patients with LEAs were extracted from the SerbVasc registry, mean age of 69.06±10.63 years. Major LEAs were performed in 59%, while minor LEAs in 41% of patients. Diabetes was seen in 65.1% of the patients, with 44% of them being on insulin therapy. Before LEA, only 20.3% of patients had previous peripheral revascularization. Soft tissue infection, irreversible acute ischemia, and Fontaine III and IV grade ischemia were the most common causes of above-the-knee amputations while diabetic foot was the most common cause of transphalangeal and toe amputations. The infection rate was 3.7%, the re-amputation rate was 5.7%, and the overall mortality rate was 6.9%, with intrahospital mortality in patients with above-the-knee amputation of 11.1%. The most significant intrahospital mortality predictors were age >65 years (p<0.001), chronic kidney disease (CKD) (p<0.001), ischemic heart disease (IHD) (p=0.001), previous myocardial revascularization (p=0.017), emergency type of admission (p<0.001), not using aspirin (p=0.041), using previous anticoagulation therapy (p=0.003), and postoperative complications (p<0.001). CONCLUSIONS: The main predictors of increased mortality after LEAs from the SerbVasc registry are age >65 years, CKD, IHD, previous myocardial revascularization, emergency type of admission, not using aspirin, using previous anticoagulation therapy, and postoperative complications. Taking into account high mortality rates after LEAs and a small proportion of previous peripheral revascularization, the work should be done on early diagnosis and timely treatment of PAD hopefully leading to decreased number of LEAs and overall mortality. CLINICAL IMPACT: Mortality after lower limb amputation from the SerbVasc register is high. A small number of previously revascularized patients is of particular clinical importance, bearing in mind that the main reasons for above-the-knee amputations were irreversible ischemia, Fontaine III and Fontaine IV grade ischemia. Lack of diagnostics procedures and late recognition of patients with PAD, led to subsequent threating limb ischemia and increased amputation rates. The work should be done on early diagnosis and timely treatment of PAD in Serbia, hopefully leading to an increased number of PAD procedures, decreased number of LEAs, and lower overall mortality.

Agreement between Lea Symbols and Patti Pics visual acuity in children and adults

Background Patti Pics (PP) and Lea Symbols (LS) are commonly used by eye care practitioners worldwide. Although the relationship between the two tests is fairly well understood, the availability of different chart designs (single optotypes, multiple optotypes, multiple optotypes with crowding box) merits futher understanding. The purpose of this study is to explore the agreement between the acuity measures obtained with Patti Pics and Lea Symbols in children and adults and compare their performance with the Sloan Letter (SL) chart in adults. Methods Monocular visual acuity was obtained from ninety-three 3 to 5-year-old children using Patti Pics and Lea Symbols. Acuities were also obtained from 113 adults using the same tests under identical conditions. Acuity results obtained with the pediatric tests were compared with the gold-standard Sloan Letter chart in adults. The Bland-Altman method was implemented to compare the level of agreement between tests. Results Patti Pics yielded worse visual acuity than the Lea Symbols by approximately half a logMAR line in both children (mean difference: -0.07 ± 0.07 logMAR, p <0.01) and adults (Mean difference: -0.05 ± 0.06 logMAR, p <0.01). The 95% limits of agreement between Lea Symbol acuity and Patti pics acuity in children was ± 0.14 logMAR. Mean difference between the Sloan Letter chart and Lea Symbols acuity was not statistically significant (p = 0.08) in adults but the difference was statistically significant between PP and SL (p<0.001). The 95% limits of agreement between LS and SL and between PP and SL was ± 0.19 logMAR and ± 0.22 logMAR, respectively. Conclusion Patti Pics consistently underestimated visual acuity as compared to Lea Symbols both in children and adults although the differences were not clinically significant. The LS and PP did not yield clinically significant differences in acuities when compared with Sloan letters in adults. (AU)

The Impact of a Short-Term Ketogenic Low-Carbohydrate High-Fat Diet on Biomarkers of Intestinal Epithelial Integrity and Gastrointestinal Symptoms.

Endurance exercise can disturb intestinal epithelial integrity, leading to increased systemic indicators of cell injury, hyperpermeability, and pathogenic translocation. However, the interaction between exercise, diet, and gastrointestinal disturbance still warrants exploration. This study examined whether a 6-day dietary intervention influenced perturbations to intestinal epithelial disruption in response to a 25-km race walk. Twenty-eight male race walkers adhered to a high carbohydrate (CHO)/energy diet (65% CHO, energy availability = 40 kcal·kg FFM-1·day-1) for 6 days prior to a Baseline 25-km race walk. Athletes were then split into three subgroups: high CHO/energy diet (n = 10); low-CHO, high-fat diet (LCHF: n = 8; <50 g/day CHO, energy availability = 40 kcal·kg FFM-1·day-1); and low energy availability (n = 10; 65% CHO, energy availability = 15 kcal·kg FFM-1·day-1) for a further 6-day dietary intervention period prior to a second 25-km race walk (Adaptation). During both trials, venous blood was collected pre-, post-, and 1 hr postexercise and analyzed for markers of intestinal epithelial disruption. Intestinal fatty acid-binding protein concentration was significantly higher (twofold increase) in response to exercise during Adaptation compared to Baseline in the LCHF group (p = .001). Similar findings were observed for soluble CD14 (p < .001) and lipopolysaccharide-binding protein (p = .003), where postexercise concentrations were higher (53% and 36%, respectively) during Adaptation than Baseline in LCHF. No differences in high CHO/energy diet or low energy availability were apparent for any blood markers assessed (p > .05). A short-term LCHF diet increased intestinal epithelial cell injury in response to a 25-km race walk. No effect of low energy availability on gastrointestinal injury or symptoms was observed.

Genome-wide identification of late embryogenesis abundant protein family and their key regulatory network in Pinus tabuliformis cold acclimation.

Cold acclimation is a crucial biological process that enables conifers to overwinter safely. The late embryogenesis abundant (LEA) protein family plays a pivotal role in enhancing freezing tolerance during this process. Despite its importance, the identification, molecular functions and regulatory networks of the LEA protein family have not been extensively studied in conifers or gymnosperms. Pinus tabuliformis, a conifer with high ecological and economic values and with high-quality genome sequence, is an ideal candidate for such studies. Here, a total of 104 LEA genes were identified from P. tabuliformis, and we renamed them according to their subfamily group: PtLEA1-PtLEA92 (group LEA1-LEA6), PtSMP1-PtSMP6 (group seed maturation protein) and PtDHN1-PtDHN6 (group Dehydrin). While the sequence structure of P. tabuliformis  LEA genes are conserved, their physicochemical properties exhibit unique characteristics within different subfamily groupings. Notably, the abundance of low-temperature responsive elements in PtLEA genes was observed. Using annual rhythm and temperature gradient transcriptome data, PtLEA22 was identified as a key gene that responds to low-temperature induction while conforming to the annual cycle of cold acclimation. Overexpression of PtLEA22 enhanced Arabidopsis freezing tolerance. Furthermore, several transcription factors potentially co-expressed with PtLEA22 were validated using yeast one-hybrid and dual-luciferase assays, revealing that PtDREB1 could directly bind PtLEA22 promoter to positively regulate its expression. These findings reveal the genome-wide characterization of P. tabuliformis  LEA genes and their importance in the cold acclimation, while providing a theoretical basis for studying the molecular mechanisms of cold acclimation in conifers.

Characterization of wheat Wrab18 gene promoter and expression analysis under abiotic stress.

BACKGROUND: Promoters play key roles in plant gene expression in complex and varied natural environments. The type and amount of cis-acting elements in the promoter sequence tend to indicate the response of genes to induction factors. WRAB18 is a group III member of the late embryogenesis abundant (LEA) protein family that performs multiple functions in plant stress physiology. To elucidate the particularly biological effects of WRAB18 on stress, exploration of its promoter sequence is necessary. METHODS AND RESULTS: In this study, the full-length and promoter sequences of Wrab18 were isolated from the Zhengyin 1 cultivar of Triticum aestivum. The gene sequences and cis-acting elements in the promoter were analyzed using the Plant Promoter Database and bioinformatics methods. The results showed that Wrab18 possessed one intron with 100 bp, the promoter sequence contained various stress-related cis-acting elements, and the functionality of the promoter was checked using green fluorescent protein (GFP) marker protein expression by transient assay in Nicotiana benthamiana. Furthermore, based on promoter prediction analysis, quantitative real-time fluorescent PCR results confirmed the response of gene expression levels to stress factors. CONCLUSIONS: In summary, the promoter sequence of Wrab18 plays a role in plant stress responses, contains multiple cis-acting elements, and provides insights into the role of WRAB18 in plant resilience to stress. This study has guiding significance for further studies of gene function and mechanism of action, and lays a theoretical foundation for improving wheat quality.