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Cereal grains are an important source of food and feed. To provide comprehensive spatiotemporal information about biological processes in developing seeds of cultivated barley (Hordeum vulgare L. subsp. vulgare), we performed a transcriptomic study of the embryo, endosperm, and seed maternal tissues collected from grains 4-32 days after pollination. Weighted gene co-expression network and motif enrichment analyses identified specific groups of genes and transcription factors (TFs) potentially regulating barley seed tissue development. We defined a set of tissue-specific marker genes and families of TFs for functional studies of the pathways controlling barley grain development. Assessing selected groups of chromatin regulators revealed that epigenetic processes are highly dynamic and likely play a major role during barley endosperm development. The repressive H3K27me3 modification is globally reduced in endosperm tissues and at specific genes related to development and storage compounds. Altogether, this atlas uncovers the complexity of developmentally regulated gene expression in developing barley grains., Competing Interests: Conflict of interest statement. The authors declare no conflict of interest. The funders had no role in the design of the study, in the collection, analyses, or interpretation of data, in the writing of the manuscript, or in the decision to publish the results., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

DNA damage response (DDR) is an essential mechanism by which living organisms maintain their genomic stability. In plants, DDR is important also for normal growth and yield. Here, we explored the DDR of a temperate model crop barley (Hordeum vulgare) at the phenotypic, physiological, and transcriptomic levels. By a series of in vitro DNA damage assays using the DNA strand break (DNA-SB) inducing agent zeocin, we showed reduced root growth and expansion of the differentiated zone to the root tip. Genome-wide transcriptional profiling of barley wild-type and plants mutated in DDR signaling kinase ATAXIA TELANGIECTASIA MUTATED AND RAD3-RELATED (hvatr.g) revealed zeocin-dependent, ATR-dependent, and zeocin-dependent/ATR-independent transcriptional responses. Transcriptional changes were scored also using the newly developed catalog of 421 barley DDR genes with the phylogenetically-resolved relationships of barley SUPRESSOR OF GAMMA 1 (SOG1) and SOG1-LIKE (SGL) genes. Zeocin caused up-regulation of specific DDR factors and down-regulation of cell cycle and histone genes, mostly in an ATR-independent manner. The ATR dependency was obvious for some factors associated with DDR during DNA replication and for many genes without an obvious connection to DDR. This provided molecular insight into the response to DNA-SB induction in the large and complex barley genome., (© 2024. The Author(s).)

Precise localization and dissection of gene promoters are key to understanding transcriptional gene regulation and to successful bioengineering applications. The core RNA polymerase II initiation machinery is highly conserved among eukaryotes, leading to a general expectation of equivalent underlying mechanisms. Still, less is known about promoters in the plant kingdom. In this study, we employed cap analysis of gene expression (CAGE) at three embryonic developmental stages in barley to accurately map, annotate, and quantify transcription initiation events. Unsupervised discovery of de novo sequence clusters grouped promoters based on characteristic initiator and position-specific core-promoter motifs. This grouping was complemented by the annotation of transcription factor binding site (TFBS) motifs. Integration with genome-wide epigenomic data sets and gene ontology (GO) enrichment analysis further delineated the chromatin environments and functional roles of genes associated with distinct promoter categories. The TATA-box presence governs all features explored, supporting the general model of two separate genomic regulatory environments. We describe the extent and implications of alternative transcription initiation events, including those that are specific to developmental stages, which can affect the protein sequence or the presence of regions that regulate translation. The generated promoterome dataset provides a valuable genomic resource for enhancing the functional annotation of the barley genome. It also offers insights into the transcriptional regulation of individual genes and presents opportunities for the informed manipulation of promoter architecture, with the aim of enhancing traits of agronomic importance., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

Rabl organization is a type of interphase chromosome arrangement with centromeres and telomeres clustering at opposite nuclear poles. Here, we analyzed nuclear morphology and chromosome organization in cycling and endoreduplicated nuclei isolated from embryo and endosperm tissues of developing barley seeds. We show that endoreduplicated nuclei have an irregular shape, less sister chromatid cohesion at 5S rDNA loci, and a reduced amount of centromeric histone CENH3. While the chromosomes of the embryo and endosperm nuclei are initially organized in Rabl configuration, the centromeres and telomeres are intermingled within the nuclear space in the endoreduplicated nuclei with an increasing endoreduplication level. Such a loss of chromosome organization suggests that Rabl configuration is introduced and further reinforced by mitotic divisions in barley cell nuclei in a tissue- and seed age-dependent manner., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Wild barley is abundant, occupying large diversity of sites, ranging from the northern mesic Mediterranean meadows to the southern xeric deserts in Israel. This is also reflected in its wide phenotypic heterogeneity. We investigated the dynamics of DNA content changes in seed tissues in ten wild barley accessions that originated from an environmental gradient in Israel. The flow cytometric measurements were done from the time shortly after pollination up to the dry seeds. We show variation in mitotic cell cycle and endoreduplication dynamics in both diploid seed tissues (represented by seed maternal tissues and embryo) and in the triploid endosperm. We found that wild barley accessions collected at harsher xeric environmental conditions produce higher proportion of endoreduplicated nuclei in endosperm tissues. Also, a comparison of wild and cultivated barley strains revealed a higher endopolyploidy level in the endosperm of wild barley, that is accompanied by temporal changes in the timing of the major developmental phases. In summary, we present a new direction of research focusing on connecting spatiotemporal patterns of endoreduplication in barley seeds and possibly buffering for stress conditions.

Seeds are complex biological systems comprising three genetically distinct tissues: embryo, endosperm, and maternal tissues (including seed coats and pericarp) nested inside one another. Cereal grains represent a special type of seeds, with the largest part formed by the endosperm, a specialized triploid tissue ensuring embryo protection and nourishment. We investigated dynamic changes in DNA content in three of the major seed tissues from the time of pollination up to the dry seed. We show that the cell cycle is under strict developmental control in different seed compartments. After an initial wave of active cell division, cells switch to endocycle and most endoreduplication events are observed in the endosperm and seed maternal tissues. Using different barley cultivars, we show that there is natural variation in the kinetics of this process. During the terminal stages of seed development, specific and selective loss of endoreduplicated nuclei occurs in the endosperm. This is accompanied by reduced stability of the nuclear genome, progressive loss of cell viability, and finally programmed cell death. In summary, our study shows that endopolyploidization and cell death are linked phenomena that frame barley grain development., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Understanding the mechanisms regulating the development of cereal seeds is essential for plant breeding and increasing yield. However, the analysis of cereal seeds is challenging owing to the minute size, the liquid character of some tissues, and the tight inter-tissue connections. Here, we demonstrate a detailed protocol for dissection of the embryo, endosperm, and seed maternal tissues at early, middle, and late stages of barley seed development. The protocol is based on a manual tissue dissection using fine-pointed tools and a binocular microscope, followed by ploidy analysis-based purity control. Seed maternal tissues and embryos are diploid, while the endosperm is triploid tissue. This allows the monitoring of sample purity using flow cytometry. Additional measurements revealed the high quality of RNA isolated from such samples and their usability for high-sensitivity analysis. In conclusion, this protocol describes how to practically dissect pure tissues from developing grains of cultivated barley and potentially also other cereals.

Aims: At specific time periods following ischemic stroke (IS), acute coronary syndrome as ischemic heart disease (IHD) represents a higher risk of death than IS. Not all IS patients can undergo specific examination for IHD detection. The aim of this study was to assess exclusive risk factors (RFs) associated with IHD occurrence in IS patients. Knowledge of these RFs should help in stratifying IS patients for IHD detection., Materials and Methods: This was a hospital-based, retrospective, single centre study. The sample consisted of 192 consecutive IS patients, divided into two subgroups - Subgroup 1 (54 patients without IHD; 55.6% males; 63.1 ± 11.8 years) and Subgroup 2 (138 patients with IHD; 39.1% males; 76.3 ± 9.6 years). The following factors were identified: age; sex; presence of arterial hypertension, atrial fibrillation, diabetes mellitus; plasma levels of total cholesterol, triglycerides, low-density cholesterol, high-density cholesterol; body mass index; presence of carotid plaques. Logistic regression analysis was used for statistical evaluation., Results: Of all identified risk factors only age (OR=1.109; 95% CI: 1.069 - 1.150, P=0.001) and the presence of arterial hypertension (OR=6.298; 95% CI: 2.215 - 17.905, P=0.003) were exclusively and significantly associated with the presence of IHD in IS patients., Conclusions: Age and arterial hypertension may be exclusive risk factors associated with IHD in IS patients.

Background: Antiplatelet therapy plays a crucial role in the primary and secondary prevention of noncardioembolic ischemic stroke / transient ischemic attacks (IS/TIA). Several antiplatelet agents are available. This review deals with the characteristics of particular antiplatelet agents as well as choice of antiplatelet treatment in various situations, based on the evidence and international recommendations., Methods: PubMed and Stroke Trials Registry on-line databases and the European Stroke Organisation Guidelines for Management of IS/TIA 2008 and update of the recommendations of the American Heart Association / American Stroke Association Council 2008 on Stroke were used., Results: Acetylsalicylic acid (ASA) is the only antiplatelet drug used in primary prevention, mainly to reduce the risk of myocardial infarction (MI), but also in women aged 45 years or more and in some patients with non-valvular atrial fibrillation to reduce risk of IS/TIA. In the secondary prevention of noncardioembolic IS/TIA, ASA in combination with long release dipyridamole (DIP) and clopidogrel (CLOP) alone are considered first choice therapies. The choice of the particular antiplatelet agent should be individualized according to the patient risk factor profiles and treatment tolerance. ASA alone or triflusal can be used alternatively in patients who cannot be treated with either ASA+DIP or CLOP. The use of indobufen should be considered only in patients in need of temporary interruption of the antiplatelet therapy. Ticlopidine (TIC) should not be newly introduced into the treatment. Currently, insufficient data are available on the use of cilostazol in IS/TIA prevention.

Robot-assisted minimally invasive surgery has become a routine surgical option for the treatment of prostate cancer. Despite its technical advancements, the da Vinci(®) Surgical System still lacks haptic feedback to the surgeon, resulting in a maximally applied compressive force by the robotic needle driver during every grasping maneuver. Without this perceptional sense of touch and grip control, repetitive robotic needle driver manipulation may unknowingly lead to irreparable damage to fine sutures used during delicate anastomotic repairs. For robotic prostatectomy, any such loss of integrity can potentially lead to premature breakdown of the urethrovesical anastomosis and urine extravasation, especially important for a less-than-perfectly fashioned anastomotic repair. Although it has already been established that overhandling of sutures using handheld laparoscopic instruments can lead to reduced suture strength, it has not been established to what extent this may occur after robotic surgical procedures. We present analytical data and analyses concerning the failure strength of fine sutures commonly used for urethrovesical anastomotic repair during robotic prostatectomy, after repetitive robotic needle driver manipulation. When compared with noncompromised monofilament suture controls, the average maximal failure force after repetitive robotic manipulation was significantly reduced by 35% (p < 0.0001). Similarly, the average maximal failure force of braided sutures was significantly reduced after repetitive robotic manipulation by 3% (p = 0.009). This work demonstrates that significant reductions in monofilament and braided suture strength integrity can occur after customary repetitive manipulation by robotic needle drivers in an ex vivo model, with further research warranted in the in vivo setting.

The most common clinical cause of long-term failure in total joint replacement surgery is inflammatory aseptic osteolysis; a condition in which bone surrounding the prosthetic implant, and to which the implant is attached, is resorbed, rendering the artificial device loose and painful. Historically, the severity of this bone resorptive process has been thought to be predominately attributed to the size and shape of wear-debris particles, particularly the metallic particulates that interact biologically/immunologically with cells in the joint. Because the cytotoxic reactions are the result of interactions between the cells and the surfaces of the particulates, it is not clear in the realm of orthopedics to what extent different surface stoichiometric ratios contribute to instigating bioreactive or cytotoxic cellular responses that can lead to aseptic osteolysis. Using energy dispersive X-ray spectroscopy (EDS) and X-ray photoelectron spectroscopy (XPS), this study presents data and analyses concerning the respective bulk and surface stoichiometric ratios of two commercially pure metal micro-particulates (tantalum and titanium), two prosthetic F75 cobalt-chromium-molybdenum alloy micro-particulates, and prosthetic F136 titanium-aluminum-vanadium alloy micro-particulates, each containing elements common to total joint replacement surgery. Cell culture viability data from four volunteer donors are also presented, which suggest that micro-particulates containing large percentages of surface titanium and aluminum can cause moderate cellular toxicity, and micro-particulates containing large percentages of surface cobalt can result in extremely severe cellular toxicity. This work further suggests that surface analysis techniques, such as XPS, are essential to determine surface elemental characterization of metallic materials prior to interpreting cellular response results.

Introduction: The assessment of methods for the prevention of postoperative pelvic adhesions is hampered by the lack of a quantifiable adhesion measurement technique. Currently available methods for adhesion model assessment rely on qualitative grading scales; a quantitative method would have many attendant benefits, including standardized reporting., Methods: A technique was developed to generate consistent and significant adhesions in a swine model that are suitable for quantitative assessment using a Material Testing System (MTS) machine platform. In this method, the uterine horns are cannulated and then attached to the pelvic sidewall using loose silk sutures. The underlying sidewall and adjacent uterine serosa are injured with electrocautery. Following a two-week survival the entire complex of uterine horn and sidewall are excised en bloc and prepared for MTS testing., Results: Extrication of the adhesion, as measured by the MTS platform, generates a quantitative assessment of adhesion strength that can be scaled and analyzed to produce several continuous variable descriptions., Discussion: This technique represents a novel quantitative method for adhesion assessment in an animal model. This quantitative technique may then be applied to accurately assess a range of adhesion prevention techniques, producing results which can be standardized for comparison.

We present data and analyses concerning the cytotoxicity and bioreactivity associated with the surface composition of fine metal particulates that are similar to those commonly released in the body by prostheses used in total joint replacement surgery. Here we study the bulk and surface compositions of three separately procured cobalt-chromium-molybdenum (CoCrMo) micron-sized particulate powders, each identified by their corresponding vendor as being ASTM F75 grade material. We use energy dispersive spectroscopy (EDS) to verify the bulk metallic composition and X-ray photoelectron spectroscopy (XPS) to examine the surface metallic composition of each CoCrMo powder. Cultured synovial fibroblasts were then exposed to the particulate powders to see how the metallic surfaces might affect cellular viability. Results indicate that while the bulk metallic composition of each CoCrMo powder was similar, the surface metallic compositions were found to be dramatically different and yielded equally dramatic differences in terms of cytotoxicity and bioreactivity of synovial fibroblast in culture.

While the industrial community already employs multiple surface analytical techniques to study compositional wearing of various metallic and nonmetallic materials, as yet, these methods have not been widely introduced into the biological community. We report on a novel approach, using the industrial spectroscopic techniques of X-ray photoelectron spectroscopy, micro-Raman spectroscopy, and scanning electron microscopy equipped with energy dispersive spectroscopy, to identify the fine wear particulates and other impurities deposited within the knee-joint following total knee arthroplasty. In this study, synovial fluid was extracted from knee-joints scheduled for revision of total knee arthroplasty. The small debris flake formed by centrifugation of the fluid was analyzed using the spectroscopic techniques mentioned above. These nondestructive techniques were successful in identifying numerous micron and submicron sized metallic particulates that appear to emanate from both the prosthetic bearing (articulating) surfaces and from backside (nonarticulating) surfaces, even when gross wearing of the prosthetic device was not detectable by direct visual inspection intraoperatively. Most interesting is that the ratio of the in vivo metallic debris is approximately the same ratio as that of the manufactured alloy, indicating prosthetic wearing as opposed to chemical dissolution. More importantly, using these spectroscopic techniques to probe both the surface and below the surface of the synovial deposits, we identify an inhomogeneous distribution of the wear debris. This indicates the need to use multiple techniques in order to adequately identify the elemental composition of the prosthetic wear material., ((c) 2007 Wiley Periodicals, Inc.)

Purpose: To present a novel in-vitro pig model for testing the design and function of different internal catheter constructs, along with the resulting exit wound resulting from removal., Materials and Methods: We compared the pullout parameters of two nephrostomy catheters that differ only in their internal loop design., Results: Greater force, work, and displacement are necessary to extricate catheters with a double loop without increasing exit-wound size., Conclusion: Although the differences were not statistically significant, results from this model have potential clinical implications that may prompt further design modifications and prototype development to improve the retention of catheters in the kidney, thus reducing patient discomfort and costs associated with repeated catheter placement.

Interleukin-1beta and tartrate resistant acid phosphatase concentrations in synovial fluid aspirates were examined to determine if they could be used as indicators of increased synovial inflammation and an osteolytic reaction in patients having total knee arthroplasty. Synovial aspirates were obtained from seven patients with severely osteoarthritic knees that were scheduled for primary total knee arthroplasty and from 20 patients with knees scheduled for total knee arthroplasty revision. Eleven of the revision cases involved titanium alloy prostheses and nine involved cobalt chrome alloy prostheses. The interleukin-1beta and tartrate resistant acid phosphatase concentrations were obtained and compared between the group having primary total knee arthroplasty and the group having revision total knee arthroplasty. The knees having revision surgery had higher concentrations of interleukin-1beta and tartrate resistant acid phosphatase than did the knees having primary total knee arthroplasty. These results indicate a greater inflammatory and osteolytic response in knees having revision surgery. Although the osteoarthritic knees and the knees needing revision surgery in this study are considered to have an inflammatory state, it was only after total knee arthroplasty when particulate wear debris would be present that appreciable concentrations of interleukin-1beta and tartrate resistant acid phosphatase were produced.

The generation of metal particles through surface wear of prosthetic joints has been associated with biological reactions that may lead to prosthetic component loosening. The role of the macrophage in these reactions has been studied extensively, but that of the fibroblast has not. The few fibroblast studies that there have been have shown that particles of several metals, with sizes over a wide range, can promote cytokine release and may cause cell necrosis. The intent of this study was to determine if there are metal particle exposure threshold levels that result in morphological changes and cell necrosis of fibroblasts in peri-articular tissues. Retrieved human fibroblasts (superior medial plica) were cultured in standard fashion and then were exposed to various particle dosages of commercially pure Titanium (cpTi). Cell morphological changes and necrosis were observed to occur when the total mass of the particle dosage exceeded a threshold level. These data imply that these cell responses occur at threshold levels of wear particle exposure., (Copyright 1999 John Wiley & Sons, Inc.)

The loosening of total joint arthroplasties (TKA) with associated osteolysis has been a persistent problem in orthopaedics. Wear debris from prosthetic devices including Titanium (Ti) is involved in this process. Mechanisms for this osteolytic process are unclear. The purpose of this study was to compare the biological response of Ti and Tantalum (Ta) on retrieved human fibroblasts. Fibroblasts were retrieved from human volunteers and cultured using standard techniques. Twenty-five (25) ml culture flasks were seeded with cells and when reaching confluency four concentrations of Ti and Ta were added. Their mean size was less than 3 microns for both metals and gram weights were 0.0048. 0.0096, 0.048, and 0.096 gms. After ten (10) days the cells were fixed, stained and photographed. For both Ti and Ta, the lowest concentration had little effect on the cells, while at the two higher concentrations, nearly all of the cell were killed. Since both of the metals tested are considered to be inert with respect to toxicity, these results would suggest that the observed cell death, seen equally for both metals, was due to the size and concentration of the particles and not to the metals tested. Mechanisms are currently being investigated which include mechanical as well as chemical factors.

Interleukin-1 beta levels from synovial aspirates were examined to determine if they can be used as indicators of increased synovial activity and an inflammatory reaction within total knee arthroplasty. Synovial aspirates were obtained from twelve osteoarthritic knees scheduled for total knee arthroplasty and twenty-one knees scheduled for total knee revision. Eleven of the revision cases involved titanium alloy prostheses and ten involved cobalt-chrome prostheses. Using a high sensitivity ELISA test kit, the interleukin-1 beta concentrations were compared. A significant difference in the interleukin-1 beta concentration was found between all three knee groups. The knees scheduled for revision surgery showed higher concentrations of interleukin-1 beta than osteoarthritic knees. While the knees implanted with a titanium prosthesis showed the greatest concentration of interleukin-1 beta, the osteoarthritic knees showed the lowest interleukin-1 beta concentration. Furthermore, significantly greater synovial aspirate volumes were obtained from the revision cases than from the osteoarthritic cases. This increased synovial activity is most likely attributed to the high concentration of the particulate wear debris produced from the prosthesis.

In this retrospective study, we compared forty-five patients who had undergone same-day bilateral total knee arthroplasties with 144 patients who had undergone unilateral total knee arthroplasty procedures. All procedures were performed by a single surgeon using a single prosthetic design between June 1994 to May 1996. The issues studied were the length of hospital stay, hospital charges, units of blood transfused, units of homologous blood transfused, and the rate and type of complications. This study also examined the usage and financial charges of extended care facilities and/or home health care following discharge from the hospital. Results show that, for patients suffering from symptomatic bilateral knee disease, the same-day bilateral total knee procedure is significantly less costly than staged bilateral procedures. The rate and type of complications seen in the same-day bilateral cases were not significantly different from complications seen in unilateral cases.

Electromyographic signals (EMG) from surface electrodes over the vastus medialis, rectus femoris and vastus lateralis were monitored during isometric knee extension for 10 TKA patients and 6 control subjects. No significant side-to-side differences in normalized EMG signals from any of the monitored muscles were found when the left and right legs of the control group were compared or when the operative and the non-operative legs of the patient group were compared. However, both the operative and the non-operative legs in the patient group differed significantly (p < 0.01) in normalized EMG from the control group. This study has shown that a muscle imbalance, possibly leading to patellar tracking problems, does not routinely exist following TKA through a medial parapatellar incision.

Two hundred eight primary total hip arthroplasties were reviewed to evaluate the effect of closed suction drainage. This review included 45 hips in which closed drains were used and 163 hips in which drains were not used. These two groups were compared for possible differences in wound problems, temperature elevations, changes in Hgb/Hct, and the need for transfusions. There was no statistically significant difference in postoperative temperatures or decrease in Hgb. However, there were four superficial wound infections in the drained group and three superficial wound infections in the non-drained group (P < .025). There were no deep infections in either group. These findings suggest closed suction drainage provides no apparent advantage in uncomplicated primary total hip arthroplasty.

One hundred seventy-one nurses had their back strength evaluated on an isokinetic lifting device and filled out an epidemiologic questionnaire. They were then followed prospectively for 2 years to determine the incidence of job-related low-back injuries. The data were analyzed to determine if the injury incidence correlated with any of the strength or epidemiologic variables collected during the original evaluation. Average peak force measured during the isokinetic lift was 63.8 kg + 13.6 kg at a lift speed of 30.5 cm/sec and 59.1 kg + 14.9 kg at a lift speed of 45.7 cm/sec. Sixteen nurses reported an occurrence of job-related low-back pain or injury during the 2-year prospective period. Discriminate statistical techniques showed that none of the strength or epidemiologic variables correlated with the incidence of pain or injury or explained significant amounts of variance when the variables were regressed on strength or work calculated from the lift force/lift height data. It was concluded that in this high risk population, in which loads are heavy and lifting postures are variable, the use of low-back strength or prior history of pain or injury are poor predictors as to subsequent low-back pain or injury.