Biosynthesis of the allelopathic alkaloid gramine in barley by a cryptic oxidative rearrangement.

The defensive alkaloid gramine not only protects barley and other grasses from insects but also negatively affects their palatability to ruminants. The key gene for gramine formation has remained elusive, hampering breeding initiatives. In this work, we report that a gene encoding cytochrome P450 monooxygenase CYP76M57, which we name AMI synthase (AMIS), enables the production of gramine in Nicotiana benthamiana, Arabidopsis thaliana, and Saccharomyces cerevisiae. We reconstituted gramine production in the gramine-free barley (Hordeum vulgare) variety Golden Promise and eliminated it from cultivar Tafeno by Cas-mediated gene editing. In vitro experiments unraveled that an unexpected cryptic oxidative rearrangement underlies this noncanonical conversion of an amino acid to a chain-shortened biogenic amine. The discovery of the genetic basis of gramine formation now permits tailor-made optimization of gramine-linked traits in barley by plant breeding.

The effect of ε-aminocaproic acid on blood product requirement, outcome and thromboelastography parameters in severely thrombocytopenic dogs.

BACKGROUND: No treatment other than platelet administration is known to protect against spontaneous hemorrhage in thrombocytopenic dogs. OBJECTIVES: Primary: determine if treatment with ε-aminocaproic acid (EACA) decreases the requirement for blood transfusions and improves outcome in dogs with severe thrombocytopenia. Secondary: find evidence of hyperfibrinolysis and determine the effect EACA administration on rapid (rTEG) and tissue plasminogen activator-spiked (tPA-rTEG) thromboelastography parameters. ANIMALS: Twenty-seven dogs with severe thrombocytopenia were treated with EACA, and data from an additional 33 were obtained from the hospital database as historical control (HC) cohort. METHODS: Single arm clinical trial with HCs. The EACA group dogs received EACA (100 mg/kg IV followed by a constant-rate infusion [CRI] of 400 mg/kg/24 hours). Thromboelastography before and during EACA infusion, hospitalization days, number of transfusions, and mortality were compared. RESULTS: No difference was found in number of transfusions per dog (median, interquartile range; 1, 0-2.5 vs 0.9, 0-2; P = .5) and hospitalization days (4, 4-6 vs 4.5, 3.75-6; P = .83) between HC and EACA groups, respectively, and no difference in survival was identified by log-rank analysis (P = .15). Maximum amplitude on both rTEG and tPA-rTEG increased after EACA administration (rTEG baseline: 23.6, 9.6-38.9; post-EACA: 27.3, 19.8-43.2; P < .001; tPA-rTEG baseline: 23, 10.9-37.2; post-EACA: 24.7, 16.7-44.8; P < .002). CONCLUSIONS AND CLINICAL IMPORTANCE: Although EACA increased clot strength, there was no effect on outcome. Treatment with EACA at this dosage cannot be recommended as a routine treatment but may be considered for dogs with severe ongoing hemorrhage.

Cranial autonomic symptoms in migraine are related to central sensitization: a prospective study of 164 migraine patients at a tertiary headache center.

BACKGROUND: Cranial autonomic symptoms (CASs) during migraine attacks are reported to be quite common regardless of ethnicity. In our previous study investigating 373 migraineurs, we found that 42.4% of them had CASs. The patients with CASs more frequently had cutaneous allodynia than did those without CASs, and we speculated that CASs were associated with central sensitization. The present study searched for substantial evidence on the relationship between CASs and central sensitization in migraine patients. METHODS: This was a prospective cross-sectional study. We studied a new independent cohort of 164 migraineurs who presented to the Tominaga Hospital Headache Center from July 2018 until December 2019. The clinical features of CASs according to the criteria in ICHD-3 (beta) were investigated. We also evaluated central sensitization based on the 25 health-related symptoms utilizing the validated central sensitization inventory (CSI), and each symptom was rated from 0 to 4 resulting a total score of 0-100. RESULTS: The mean age was 41.8 (range: 20 to 77) years old. One hundred and thirty-one patients (78.9%) were women. Eighty-six of the 164 (52.4%) patients had at least 1 cranial autonomic symptom. The CSI score of the patients with ≥3 CASs reflected a moderate severity and was significantly higher than in those without CASs (41.9 vs. 30.7, p = 0.0005). The score of the patients with ≥1 conspicuous CAS also reflected a moderate severity and was significantly higher than in those without CASs (40.7 vs. 33.2, p = 0.013). The patients in the CSI ≥40 group had lacrimation, aural fullness, nasal blockage, and rhinorrhea, which are cranial autonomic parasympathetic symptoms, significantly more frequently than those in the CSI < 40 group. CONCLUSIONS: Migraine patients with CASs showed significantly greater central sensitization than those without such symptoms. In particular, cranial parasympathetic symptoms were more frequent in centrally sensitized patients than in nonsensitized patients, suggesting that cranial parasympathetic activation may contribute to the maintenance of central sensitization. TRIAL REGISTRATION: This study was retrospectively registered with UMIN-CTR on 29 Aug 2020 ( UMIN000041603 ).

Hemiplegic migraine type 2 with new mutation of the ATP1A2 gene in Japanese cases.

We analyzed the clinical symptoms of hemiplegic migraine (HM) and their relevance in four Japanese patients considered to have ATP1A2 mutations as a cause. Sequencing of ATP1A2 was performed using the Sanger method in 43 blood samples from clinically suspected patients with familial HM. Subsequently, algorithm analysis, allele frequency determination, and three-dimensional structure analysis of the recognized variants were performed, and the recognized variants were evaluated. We found four heterozygous missense mutations in ATP1A2 (Case 1: p.R51C; Case 2: p.R65L; Case 3: p.A269P; Case 4: p.D999H), three of which had not been reported to date. These four mutations may also affect the structure of the protein products, as assessed using a three-dimensional structural analysis. In all four cases, the clinical symptoms included visual, sensory, motor, and verbal symptoms and the frequency and duration of headache attacks varied. Additionally, oral administration of a combination of lomerizine hydrochloride and topiramate had a partial effect in three cases. We report four missense mutations in ATP1A2. This report will be useful for the future analysis of mutations and clinical types in Asians, as well as Westerners, with migraine.

Evaluation of the changes in human milk lipid composition and conformational state with Raman spectroscopy during a breastfeed.

Human milk fat forms the main energy source for breastfed infants, and is highly variable in terms of concentration and composition. Understanding the changes in human milk lipid composition and conformational state during a breastfeed can provide insight into lipid synthesis and secretion in the mammary gland. Therefore, the aim of this study was to evaluate human milk fatty acid length, degree of unsaturation (lipid composition) and lipid phase (lipid conformational state) at different stages during a single breastfeed (fore-, bulk- and hindmilk). A total of 48 samples from 16 lactating subjects were investigated with confocal Raman spectroscopy. We did not observe any significant changes in lipid composition between fore-, bulk and hindmilk. A new finding from this study is that lipid conformational state at room temperature changed significantly during a breastfeed, from almost crystalline to almost liquid. This observation suggests that lipid synthesis in the mammary gland changes during a single breastfeed.

Ultrasound-guided paravertebral perineural glucocorticoid injection for signs of refractory cervical pain associated with foraminal intervertebral disk protrusion in four dogs.

CASE DESCRIPTION: 4 dogs, 7.5 to 10 years of age, were presented for evaluation of signs of chronic cervical pain and forelimb lameness secondary to cervical foraminal intervertebral disk protrusion (IVDP). All dogs were refractory to ≥ 2 weeks of conservative management including strict rest and pain management with anti-inflammatory drugs, methocarbamol, and gabapentin. CLINICAL FINDINGS: The MRI findings included left foraminal IVDP at C2-3 causing mild C3 nerve root compression (dog 1), multifocal degenerative disk disease with mild focal left-sided disk protrusion at C6-7 without associated spinal cord or nerve root compression (dog 2), left foraminal C6-7 IVDP with suspected focal spinal cord atrophy or mild compression (dog 3), and right foraminal C6-7 IVDP and multifocal cervical intervertebral disk degeneration with annulus fibrosus protrusion (dog 4). TREATMENT AND OUTCOME: Ultrasound-guided paravertebral perineural injections with methylprednisolone acetate (1 mg/kg [0.45 mg/lb]) at the C3 nerve root in dog 1 and at the C7 nerve root in the other 3 dogs were performed. Injections were repeated at intervals of 4 weeks to 3 months on the basis of clinical response. None of the dogs had any complications from the procedures. For dogs 1 and 4, there was complete resolution of lameness and signs of cervical pain following perineural injections, and for dog 3, there was complete resolution of lameness and only minimal residual cervical pain. Dog 2 did not have long-lasting improvement. CLINICAL RELEVANCE: Findings indicated that ultrasound-guided paravertebral perineural injection can be an effective treatment of cervical foraminal IVDP for some dogs. Additional studies to determine appropriate case selection and better assess the overall success rate and risks associated with this technique are warranted.

Enhancement of CRISPR-Cas9 induced precise gene editing by targeting histone H2A-K15 ubiquitination.

BACKGROUND: Precise genetic modifications are preferred products of CRISPR-Cas9 mediated gene editing in mammalian cells but require the repair of induced double-strand breaks (DSB) through homology directed repair (HDR). Since HDR competes with the prevailing non-homologous end joining (NHEJ) pathway and depends on the presence of repair templates its efficiency is often limited and demands optimized methodology. RESULTS: For the enhancement of HDR we redirect the DSB repair pathway choice by targeting the Ubiquitin mark for damaged chromatin at Histone H2A-K15. We used fusions of the Ubiquitin binding domain (UBD) of Rad18 or RNF169 with BRCA1 to promote HDR initiation and UBD fusions with DNA binding domains to attract donor templates and facilitate HDR processing. Using a traffic light reporter system in human HEK293 cells we found that the coexpression of both types of UBD fusion proteins promotes HDR, reduces NHEJ and shifts the HDR/NHEJ balance up to 6-fold. The HDR enhancing effect of UBD fusion proteins was confirmed at multiple endogenous loci. CONCLUSIONS: Our findings provide a novel efficient approach to promote precise gene editing in human cells.

Cranial Autonomic Symptoms of Migraine in Japan: Prospective Study of 373 Migraine Patients at a Tertiary Headache Center.

OBJECTIVE: To clarify the detailed clinical characteristics of cranial autonomic symptoms (CAS) of Japanese patients with migraine and to get insight into the pathophysiological implications. BACKGROUND: Recent studies reported that CAS in migraine is causing diagnostic confusion with cluster headache or sinus headache; however, most reports have concerned Caucasians, and Asian data are scarce. The regional differences in the clinical characteristics of primary headaches between Caucasians and Asians have also been revealed recently. METHOD: This was a cross-sectional study. We investigated 373 patients with migraine in a tertiary headache center with face-to-face interviews. RESULTS: According to our findings, 158/373 (42.4%) patients with migraine had CAS and were characterized by more frequent cutaneous allodynia than those without CAS, suggesting the contribution of central sensitization; however, there were no statistically significant differences in pulsating pain or motion sensitivity as signs of peripheral sensitization. In contrast to the previous study, osmophobia was found to be significantly related to CAS. CONCLUSION: CAS in patients with migraine is common not only in Caucasians but also in Asians. Central sensitization seems to contribute more than peripheral sensitization to CAS manifestation, and osmophobia might be noteworthy among Asian patients with migraine. To avoid a misdiagnosis, we emphasize the need for comments on CAS in the international classification of headache disorders migraine criteria.

Trauma-induced primary hypoparathyroidism following severe bite wound injury to the neck in a dog.

OBJECTIVE: To describe the clinical presentation and outcome of a dog with primary hypoparathyroidism secondary to cervical bite wounds. CASE SUMMARY: A 3-year-old male intact Chihuahua presented after being attacked by a large breed dog. The dog sustained severe cervical lacerations, exposing the trachea and jugular veins. A portion of the right thyroid gland was missing. The dog was stabilized before wound debridement and closure. Ionized calcium concentrations were within reference range at the time of presentation. Forty-eight hours after the initial trauma, the dog was presented in lateral recumbency with signs of hypovolemic shock, muscle tremors, and hyperthermia. Bloodwork showed severe ionized hypocalcemia with low normal parathyroid hormone concentration consistent with acute primary hypoparathyroidism. The dog was managed initially with IV calcium gluconate and calcitriol, then long-term oral calcium carbonate and vitamin D3. After 6 months, the dog was successfully weaned off calcium supplementation. NEW OR UNIQUE INFORMATION PROVIDED: This is the first described case of traumatic primary hypoparathyroidism after a bite injury to the neck in a dog.

The relation between emotion regulation and migraine: A cross-cultural study on the moderating effect of culture.

BACKGROUND: Effects of emotion suppression on physical health might be contingent on culture. Existing research on emotion regulation has mainly included western participants. Herewith the question arises, whether this gained expertise is transferable to an Asian culture. OBJECTIVES: This cross-sectional study evaluated to what extent the regulation of emotions is related to migraine and if the relation between emotion regulation and migraine complaints differs between a Western and an Asian population. Therefore, the main characteristics and symptoms of patients with migraine from both Germany and Japan are compared. METHODS: 261 Japanese and 347 German headache patients participated in this online study and completed self-report measures of emotion regulation (suppression and reappraisal) and headache complaints. RESULTS: Cultural groups did not differ regarding their demographic data, intake of medication and number of days with headache. German participants showed significantly higher levels of anxiety and lower levels of emotion suppression compared to Japanese patients. Emotion regulation is not correlated with headache complaints either in the Japanese or in the German patient group. CONCLUSION: Although group differences were found with respect to anxiety and emotion suppression, subsequent regression analysis revealed these differences were unrelated to headache complaints. As our baseline analysis focused on group means, approaches that examine individual reaction patterns to stress and accompanying sensory stimulus processing may prove to be more fruitful and illuminating.

Raw Cow's Milk Reduces Allergic Symptoms in a Murine Model for Food Allergy-A Potential Role For Epigenetic Modifications.

Epidemiological studies identified raw cow's milk consumption as an important environmental exposure that prevents allergic diseases. In the present study, we investigated whether raw cow's milk has the capacity to induce tolerance to an unrelated, non-milk, food allergen. Histone acetylation of T cell genes was investigated to assess potential epigenetic regulation. Female C3H/HeOuJ mice were sensitized and challenged to ovalbumin. Prior to sensitization, the mice were treated with raw milk, processed milk, or phosphate-buffered saline for eight days. Allergic symptoms were assessed after challenge and histone modifications in T cell-related genes of splenocyte-derived CD4+ T cells and the mesenteric lymph nodes were analyzed after milk exposure and after challenge. Unlike processed milk, raw milk decreased allergic symptoms. After raw milk exposure, histone acetylation of Th1-, Th2-, and regulatory T cell-related genes of splenocyte-derived CD4+ T cells was higher than after processed milk exposure. After allergy induction, this general immune stimulation was resolved and histone acetylation of Th2 genes was lower when compared to processed milk. Raw milk reduces allergic symptoms to an unrelated, non-milk, food allergen in a murine model for food allergy. The activation of T cell-related genes could be responsible for the observed tolerance induction, which suggested that epigenetic modifications contribute to the allergy-protective effect of raw milk.

Dual-comb spectroscopy using plasmon-enhanced-waveguide dispersion-compensated quantum cascade lasers.

In this Letter, we report on sub-millisecond response time mid-infrared dual-comb spectroscopy using a balanced asymmetric (dispersive) dual-comb setup with a matched pair of plasmon-enhanced-waveguide dispersion-compensated quantum cascade lasers. The system performance is demonstrated by measuring spectra of Bromomethane (CH3Br) and Freon 134a (CH2FCF3) at approximately 7.8 µm. A purely computational phase and timing-correction procedure is used to validate the coherence of the quantum cascade lasers frequency combs and to enable coherent averaging over the time scales investigated. The system achieves a noise-equivalent absorption better than 1×10-3 Hz-1/2, with a resolution of 9.8 GHz (0.326 cm-1) and an optical bandwidth of 1 THz (32 cm-1), with an average optical power of more than 1 mW per spectral element.

Histone modifications and their role in epigenetics of atopy and allergic diseases.

This review covers basic aspects of histone modification and the role of posttranslational histone modifications in the development of allergic diseases, including the immune mechanisms underlying this development. Together with DNA methylation, histone modifications (including histone acetylation, methylation, phosphorylation, ubiquitination, etc.) represent the classical epigenetic mechanisms. However, much less attention has been given to histone modifications than to DNA methylation in the context of allergy. A systematic review of the literature was undertaken to provide an unbiased and comprehensive update on the involvement of histone modifications in allergy and the mechanisms underlying this development. In addition to covering the growing interest in the contribution of histone modifications in regulating the development of allergic diseases, this review summarizes some of the evidence supporting this contribution. There are at least two levels at which the role of histone modifications is manifested. One is the regulation of cells that contribute to the allergic inflammation (T cells and macrophages) and those that participate in airway remodeling [(myo-) fibroblasts]. The other is the direct association between histone modifications and allergic phenotypes. Inhibitors of histone-modifying enzymes may potentially be used as anti-allergic drugs. Furthermore, epigenetic patterns may provide novel tools in the diagnosis of allergic disorders.

Acquired Sleep-Related Hypermotor Epilepsy with Disrupted White Matter Tracts Assessed by Multishell Diffusion Magnetic Resonance Imaging.

Sleep-related hypermotor epilepsy (SHE) (previously frontal lobe epilepsy) is a rare seizure disorder commonly misdiagnosed or unrecognized, causing negative patient sequelae. While usually reported in familial studies, it is more commonly acquired. Diagnosis is a challenge due to its low incidence in comparison with the more common sleep disorders or psychogenic etiologies in the differential diagnosis. Diagnosis is scaled on degree of certainty based on described or clinically documented semiology, with video EEG as a helpful, but not necessary, adjunct. Current treatment is similar to other focal epilepsies. We studied a 36-year-old active duty male soldier who presented with 2 years of predominantly sleep related, abrupt, short, and anamnestic hyperkinetic movements with unstructured vocalizations. Prior workup included non-contributory video electroencephalograph (EEG) and polysomnography as well as normal brain magnetic resonance imaging (MRI). Treatments for presumed psychiatric and parasomnia disturbances were not effective in establishing diagnosis or relief. Evaluation at our tertiary, multidisciplinary care institution recorded events consistent with the diagnosis of clinical SHE. He was enrolled in an advanced multishell diffusion-weighted imaging MRI research study to evaluate white matter tracts, given his history of mild, repetitive, non-penetrating traumatic brain injury, not otherwise requiring hospitalization. Multishell diffusion MRI tractography found changes not previously described in the right frontal lobe white matter tracts. These changes were consistent with neurological localization and serve as a potential nidus for this patient's seizure disorder. Misdiagnosis of SHE can result in detrimental biopsychosocial sequelae of untreated epilepsy, unnecessary or harmful intervention, or the stigmata of a behavioral disorder. Further investigation into diagnosis and etiology of acquired SHE is needed. Assessment for white matter abnormalities can potentially provide information into pathogenesis of epilepsy disorders.

3.36 µm single-mode quantum cascade laser with a dissipation below 250 mW.

We present 3.36 µm buried heterostructure distributed-feedback quantum cascade lasers with a power dissipation at threshold below 250 mW and operation temperatures as high as 130 °C. Threshold values below 20 mA at -10 °C in pulsed operation and 30 mA at -20 °C in continuous-wave operation are reported. Optical power above 130 mW and 13 mW are achieved at -20 °C in pulsed and continuous-wave operation, respectively. Continuous-wave operation occurs until 15 °C. We show single-mode emission in pulsed and continuous-wave operation. Single-mode performance is demonstrated in long pulse (5.56 µs) operation. The laser far-field exhibits a single lobe emission with full-width-half-max of 27 ° × 34 °.

Extended tuning of mid-ir quantum cascade lasers using integrated resistive heaters.

We present single mode quantum cascade lasers including a microscopic heater for spectral emission tuning. Through the use of a buried heater element, the active region temperature can be modified without changing the submount one. Emission frequency tuning in continuous-wave as large as 9 cm(-1) at 1270 cm(-1) and 14 cm(-1) at 2040 cm(-1) are observed, corresponding to an increase of the active region temperatures of ∼ 90 K. Due to the proximity of the heaters to the active region, emission can be modulated at several kHz range and the absence of moving parts guarantees the mechanical stability of the system. This method can be successfully applied to all buried heterostructure lasers, becoming an attractive solution for molecular spectroscopy in the IR. Using the presented devices, molecular absorptions of N(2)O have been measured between 1270 cm(-1) and 1280 cm(-1) and are in agreement with data from the HITRAN database.

MIDA: A Multimodal Imaging-Based Detailed Anatomical Model of the Human Head and Neck.

Computational modeling and simulations are increasingly being used to complement experimental testing for analysis of safety and efficacy of medical devices. Multiple voxel- and surface-based whole- and partial-body models have been proposed in the literature, typically with spatial resolution in the range of 1-2 mm and with 10-50 different tissue types resolved. We have developed a multimodal imaging-based detailed anatomical model of the human head and neck, named "MIDA". The model was obtained by integrating three different magnetic resonance imaging (MRI) modalities, the parameters of which were tailored to enhance the signals of specific tissues: i) structural T1- and T2-weighted MRIs; a specific heavily T2-weighted MRI slab with high nerve contrast optimized to enhance the structures of the ear and eye; ii) magnetic resonance angiography (MRA) data to image the vasculature, and iii) diffusion tensor imaging (DTI) to obtain information on anisotropy and fiber orientation. The unique multimodal high-resolution approach allowed resolving 153 structures, including several distinct muscles, bones and skull layers, arteries and veins, nerves, as well as salivary glands. The model offers also a detailed characterization of eyes, ears, and deep brain structures. A special automatic atlas-based segmentation procedure was adopted to include a detailed map of the nuclei of the thalamus and midbrain into the head model. The suitability of the model to simulations involving different numerical methods, discretization approaches, as well as DTI-based tensorial electrical conductivity, was examined in a case-study, in which the electric field was generated by transcranial alternating current stimulation. The voxel- and the surface-based versions of the models are freely available to the scientific community.

Impact of interface roughness distributions on the operation of quantum cascade lasers.

We study the impact of interface roughness on the operation of mid-IR and THz quantum cascade lasers. Particular emphasis is given towards the differences between the Gaussian and exponential roughness distribution functions, for which we present results from simulation packages based on nonequilibrium Green's functions and density matrices. The Gaussian distribution suppresses scattering at high momentum transfer which enhances the lifetime of the upper laser level in mid-IR lasers. For THz lasers, a broader range of scattering transitions is of relevance, which is sensitive to the entire profile of the interface fluctuations. Furthermore we discuss the implementation of interface roughness within a two band model.

Distributed-feedback quantum cascade laser emitting at 3.2 µm.

In this work, we present GaInAs/AlAs/AlInAs quantum cascade lasers emitting from 3.2 to 3.4 µm. Single-mode emission is obtained using buried distributed-feedback gratings fabricated using standard deep-UV contact lithography. This technique can easily be transferred to industrial production. Devices with single-mode emission down to 3.19 µm were achieved with peak power of up to 250 mW at -20 °C. A tuning range of 11 cm(-1) was obtained by changing the device temperature between -30 °C and 20 °C.

The diagnostic relevance of NT-proBNP and proANP 31-67 measurements in staging of myxomatous mitral valve disease in dogs.

BACKGROUND: There is no agreement in current publications regarding the reliability of serum concentrations of natriuretic peptides (NPs) to detect dogs with subclinical myxomatous mitral valve disease (MMVD) and to differentiate between asymptomatic stages. OBJECTIVES: We sought to compare N-terminal pro-B-type natriuretic peptide (NT-proBNP) and pro-atrial natriuretic peptide 31-67 (proANP) concentrations between various stages of canine MMVD and to investigate the influence of age, weight, and sex. METHODS: In this prospective study, dogs were classified in different disease stages using the modified Canine Heart failure International Expert Forum (CHIEF) system. Serum NP concentrations were compared between groups. RESULTS: A total of 559 samples from 116 healthy dogs and 236 dogs with MMVD were analyzed. Using cut-off values (1207 pmol/L for NT-proBNP, 1578 fmol/mL for proANP), dogs with MMVD with and without congestive heart failure (CHF) could be differentiated with a sensitivity of 83% for both and specificities of 85% and 86%, respectively. Dogs staged in CHIEF B1 and B2 could not be distinguished based on NP concentrations due to wide variation within the groups. Intact females (means 598 pmol/L and 1036 fmol/mL, respectively) had significantly higher values of both NPs than intact males (315 pmol/L and 836 fmol/mL). CONCLUSIONS: NPs in canine MMVD are useful to discriminate between asymptomatic dogs and dogs with CHF. Due to a large overlap of NP-concentrations between the groups, NPs do not seem to be useful to differentiate between dogs in stages B1 and B2. Interpretation of NT-proBNP and proANP values should include consideration of sex-specific differences.