miRNA Profiling and Its Role in Multi-Omics Regulatory Networks Connected with Somaclonal Variation in Cucumber (Cucumis sativus L.).

The role of miRNAs in connection with the phenomenon of somaclonal variation, which occurs during plant in vitro culture, remains uncertain. This study aims to investigate the possible role of miRNAs in multi-omics regulatory pathways in cucumber somaclonal lines. For this purpose, we performed sRNA sequencing (sRNA-seq) from cucumber fruit samples identified 8, 10 and 44 miRNAs that are differentially expressed between somaclones (S1, S2, S3 lines) and the reference B10 line of Cucumis sativus. For miRNA identification, we use ShortStack software designed to filter miRNAs from sRNAs according to specific program criteria. The identification of predicted in-silico targets revealed 2,886 mRNAs encoded by 644 genes. The functional annotation of miRNA's target genes and gene ontology classification revealed their association with metabolic processes, response to stress, multicellular organism development, biosynthetic process and catalytic activity. We checked with bioinformatic analyses for possible interactions at the level of target proteins, differentially expressed genes (DEGs) and genes affected by genomic polymorphisms. We assume that miRNAs can indirectly influence molecular networks and play a role in many different regulatory pathways, leading to somaclonal variation. This regulation is supposed to occur through the process of the target gene cleavage or translation inhibition, which in turn affects the proteome, as we have shown in the example of molecular networks. This is a new approach combining levels from DNA-seq through mRNA-seq, sRNA-seq and in silico PPI in the area of plants' somaclonal variation.

Profiling the Proteome of Cyst Nematode-Induced Syncytia on Tomato Roots.

Cyst nematodes are important herbivorous pests in agriculture that obtain nutrients through specialized root structures termed syncytia. Syncytium initiation, development, and functioning are a research focus because syncytia are the primary interface for molecular interactions between the host plant and parasite. The small size and complex development (over approximately two weeks) of syncytia hinder precise analyses, therefore most studies have analyzed the transcriptome of infested whole-root systems or syncytia-containing root segments. Here, we describe an effective procedure to microdissect syncytia induced by Globodera rostochiensis from tomato roots and to analyze the syncytial proteome using mass spectrometry. As little as 15 mm2 of 10-µm-thick sections dissected from 30 syncytia enabled the identification of 100-200 proteins in each sample, indicating that mass-spectrometric methods currently in use achieved acceptable sensitivity for proteome profiling of microscopic samples of plant tissues (approximately 100 µg). Among the identified proteins, 48 were specifically detected in syncytia and 7 in uninfected roots. The occurrence of approximately 50% of these proteins in syncytia was not correlated with transcript abundance estimated by quantitative reverse-transcription PCR analysis. The functional categories of these proteins confirmed that protein turnover, stress responses, and intracellular trafficking are important components of the proteome dynamics of developing syncytia.

Surgical release of the pediatric trigger thumb.

PURPOSE: The spontaneous recovery rate for locked pediatric trigger thumb (PTT) has recently been reported at between 24% and 66%; these studies concluded that a conservative approach for this condition could be adopted. The aims of this study were to review our results of surgical release of the PTT and to survey pediatric hand surgeons regarding their practice patterns for treatment of the PTT. METHODS: After institutional review board approval, we retrospectively reviewed 173 consecutive patients with 217 thumbs treated surgically at our institution. An e-mail survey of 27 pediatric hand surgeons questioned treatment of a 2-year-old child with a 6-month history of a locked trigger thumb and of an intermittently triggering thumb. RESULTS: The retrospective review demonstrated that preoperative range of motion averaged 36° loss of extension (range, 0° to 90°; SD, 22°); postoperative range of motion averaged 1° loss of extension (range, 0° to 30°; SD, 7°) at 27-day follow-up. Using a parent questionnaire at an average follow-up of 4.2 years, there were no major complications or recurrences identified. Five thumbs developed minor skin complications that healed with conservative management. There were no secondary surgeries. The practice pattern survey demonstrated that 85% of pediatric hand surgeons would treat a locked PTT in a 2-year-old with surgical release and 52% would treat an intermittently triggering thumb in a 2-year-old with continued observation if the triggering thumb was not painful. CONCLUSIONS: The surgical results reported in this study, along with the practice pattern survey, confirm that surgical release is a short, safe, and effective procedure when performed by specialty trained hand surgeons, and it is the treatment of choice for a locked PTT.

Micro MRI of the mouse brain using a novel 400 MHz cryogenic quadrature RF probe.

The increasing number of mouse models of human disease used in biomedical research applications has led to an enhanced interest in non-invasive imaging of mice, e.g. using MRI for phenotyping. However, MRI of small rodents puts high demands on the sensitivity of data acquisition. This requirement can be addressed by using cryogenic radio-frequency (RF) detection devices. The aim of this work was to investigate the in vivo performance of a 400 MHz cryogenic transmit/receive RF probe (CryoProbe) designed for MRI of the mouse brain. To characterize this novel probe, MR data sets were acquired with both the CryoProbe and a matched conventional receive-only surface coil operating at room temperature (RT) using conventional acquisition protocols (gradient and spin echo) with identical parameter settings. Quantitative comparisons in phantom and in vivo experiments revealed gains in the signal-to-noise ratio (SNR) of 2.4 and 2.5, respectively. The increased sensitivity of the CryoProbe was invested to enhance the image quality of high resolution structural images acquired in scan times compatible with routine operation (< 45 min). In high resolution (30 x 30 x 300 microm(3)) structural images of the mouse cerebellum, anatomical details such as Purkinje cell and molecular layers could be identified. Similarly, isotropic (60 x 60 x 60 microm(3)) imaging of mouse cortical and subcortical areas revealed anatomical structures smaller than 100 microm. Finally, 3D MR angiography (52 x 80 x 80 microm(3)) of the brain vasculature enabled the detailed reconstruction of intracranial vessels (anterior and middle cerebral artery). In conclusion, this low temperature detection device represents an attractive option to increase the performance of small animal MR systems operating at 9.4 Tesla.

Case report: reconstruction of a recalcitrant scapular neck nonunion and literature review.

We present the first reported treatment failure of a reconstructed scapula body that proceeded to nonunion. This is a unique case report of an otherwise healthy patient who underwent open reduction and internal fixation of a scapula fracture nonunion, which is very rare. Failure of internal fixation in this application has not been reported, and, to our knowledge, this is only the fifth case report of a scapula body nonunion that was reconstructed. Of 159 reported cases of open reduction and internal fixation for treatment of scapula neck and body fractures (with or without intraarticular glenoid fractures), there is not one reported case of a nonunion. Our case is described in detail, including the method of surgical reconstruction, and a review of the literature regarding surgical treatment of scapula nonunions after nonoperative treatment also is presented.

The suppression of tomato defence response genes upon potato cyst nematode infection indicates a key regulatory role of miRNAs.

Potato cyst nematode Globodera rostochiensis is an obligate parasite of solanaceous plants, triggering metabolic and morphological changes in roots which may result in substantial crop yield losses. Previously, we used the cDNA-AFLP to study the transcriptional dynamics in nematode infected tomato roots. Now, we present the rescreening of already published, upregulated transcript-derived fragment dataset using the most current tomato transcriptome sequences. Our reanalysis allowed to add 54 novel genes to 135, already found as upregulated in tomato roots upon G. rostochiensis infection (in total - 189). We also created completely new catalogue of downregulated sequences leading to the discovery of 76 novel genes. Functional classification of candidates showed that the 'wound, stress and defence response' category was enriched in the downregulated genes. We confirmed the transcriptional dynamics of six genes by qRT-PCR. To place our results in a broader context, we compared the tomato data with Arabidopsis thaliana, revealing similar proportions of upregulated and downregulated genes as well as similar enrichment of defence related transcripts in the downregulated group. Since transcript suppression is quite common in plant-nematode interactions, we assessed the possibility of miRNA-mediated inverse correlation on several tomato sequences belonging to NB-LRR and receptor-like kinase families. The qRT-PCR of miRNAs and putative target transcripts showed an opposite expression pattern in 9 cases. These results together with in silico analyses of potential miRNA targeting to the full repertoire of tomato R-genes show that miRNA mediated gene suppression may be a key regulatory mechanism during nematode parasitism.

Enhanced Fluorine-19 MRI Sensitivity using a Cryogenic Radiofrequency Probe: Technical Developments and Ex Vivo Demonstration in a Mouse Model of Neuroinflammation.

Neuroinflammation can be monitored using fluorine-19 (19F)-containing nanoparticles and 19F MRI. Previously we studied neuroinflammation in experimental autoimmune encephalomyelitis (EAE) using room temperature (RT) 19F radiofrequency (RF) coils and low spatial resolution 19F MRI to overcome constraints in signal-to-noise ratio (SNR). This yielded an approximate localization of inflammatory lesions. Here we used a new 19F transceive cryogenic quadrature RF probe ( 19 F-CRP) that provides the SNR necessary to acquire superior spatially-resolved 19F MRI. First we characterized the signal-transmission profile of the 19 F-CRP. The 19 F-CRP was then benchmarked against a RT 19F/1H RF coil. For SNR comparison we used reference compounds including 19F-nanoparticles and ex vivo brains from EAE mice administered with 19F-nanoparticles. The transmit/receive profile of the 19 F-CRP diminished with increasing distance from the surface. This was counterbalanced by a substantial SNR gain compared to the RT coil. Intraparenchymal inflammation in the ex vivo EAE brains was more sharply defined when using 150 µm isotropic resolution with the 19 F-CRP, and reflected the known distribution of EAE histopathology. At this spatial resolution, most 19F signals were undetectable using the RT coil. The 19 F-CRP is a valuable tool that will allow us to study neuroinflammation with greater detail in future in vivo studies.

Operative treatment of acute distal femur fractures: systematic review of 2 comparative studies and 45 case series (1989 to 2005).

BACKGROUND: The incidence of distal femur fractures is approximately 37 per 100,000 person-years. Typically, distal femur fractures are caused by a high-energy injury mechanism in young men or a low-energy mechanism in elderly women. Managing these fractures can be a challenging task. Most surgeons agree that distal femur fractures need to be treated operatively to achieve optimal patient outcomes. The articular fracture component is usually treated with open reduction and internal lag screw fixation or external tension wire fixation (Illizarov). However, there is no consensus on the type of implant for the fixation of the metaphyseal-diaphyseal fracture component. OBJECTIVE: The aim of this study is to systematically summarize and compare the results of different fixation techniques (traditional compression plating, antegrade nailing, retrograde nailing, submuscular locked internal fixation, and external fixation) in the operative management of acute nonperiprosthetic distal femur fractures (AO/OTA type 33A and C) and the characteristics of the fractures for each treatment (articular/nonarticular and open/closed). Additionally an attempt was made to evaluate the impact of surgical experience on nonunion rate, fixation failure rate, deep infection rate, and secondary surgical procedure rate. In the context of this article compression plating relates to techniques/implants that require compression of the implant to the femoral shaft-it does not relate to interfragmentary compression.

The application of dermatotraction for primary skin closure.

Management of an open wound is a problem frequently encountered in the treatment of fractures. Skin grafting, rotational flaps, free flaps, and healing by secondary intention add a considerable amount of morbidity and cost to the patient. Therefore, it is ideal to obtain primary closure when possible. This communication describes a technique that uses spinal needles, using towel clips and the natural stretching ability of the skin to enable primary closure of wounds. The technique described uses dermatotraction to stretch the skin is a cost-effective way to achieve primary closure of large wounds with supplies that are readily available in every operating room.

Mapping of scapular fractures with three-dimensional computed tomography.

BACKGROUND: Fractures of the scapula involve a unique and challenging set of considerations, which must be understood to provide optimal treatment. The primary goal of this study was to create a frequency map of a series of surgically treated scapular fractures that specifically involved the scapular body and/or neck. METHODS: A prospective database was used in the collection of consecutive radiographic imaging studies of patients undergoing operative treatment of scapular fractures. Scanned three-dimensional computed tomography images were superimposed and oriented to fit a model scapular template. Size dimensions were normalized by aligning specific scapular landmarks. Fracture lines were identified and traced over the combined three-dimensional computed tomography model to create a scapular fracture map. RESULTS: Of ninety fractures that met the criteria for inclusion, 68% involved the inferior aspect of the glenoid neck and 71% involved the superior vertebral border. Seventeen percent of the patterns included articular extension, and 22% of the fractures entered the spinoglenoid notch. Of fractures involving the inferior aspect of the glenoid neck at the lateral scapular border, 84% traversed medially to exit just inferior to the medial extent of the scapular spine, and 59% of these inferior neck fractures also had propagation to the inferior third of the vertebral border. Among the fractures involving the spinoglenoid notch, the most common pattern was demonstrated by coexisting fracture lines; 60% of the fractures of the spinoglenoid notch exited just inferior to the glenoid, 65% extended to the superior-medial vertebral border, and 45% extended to the inferior-medial vertebral border. In contrast, articular fractures did not follow predictable patterns; they demonstrated the greatest variability in trajectory, which was almost random, and there was a wide distribution of exit points along the vertebral border. CONCLUSIONS: Surgically treated scapular fractures display very common patterns. The most common pattern is the lateral border fracture immediately inferior to the glenoid, which extends to the superior vertebral border in more than two-thirds of cases. A smaller proportion of scapular fractures enter the spinoglenoid notch or the articular surface. There is great variation in the patterns of fractures involving the articular surface.

Performance of a 200-MHz cryogenic RF probe designed for MRI and MRS of the murine brain.

A cryogenic 200-MHz RF transmit/receive probe (CryoProbe) made of copper and designed for murine brain studies is described. The probe operates at 30K and its performance was compared to a home-made surface coil of equal dimensions and a mouse head volume resonator, both operating at room temperature (RT). Since for small volumes of tissue interrogated (<5 mL) sample noise and thermal noise of the receiver are of comparable magnitude, considerable noise reduction can be achieved by cooling of the receiver system, e.g., using cold helium gas. The effect of temperature lowering on the signal-to-noise ratio (SNR) was assessed in phantom and in vivo murine brain studies using conventional MRI sequences (spin and gradient echo) and single voxel MR spectroscopy. Consistent sensitivity increases were observed for the CryoProbe. SNR increases by a factor>or=2 for the phantom experiments and by factors of 1.8 to 1.9 for in vivo experiments when compared to the RT surface coil for both MRI and MRS experiments. These results are in good agreement with estimated SNR gains for a circular surface coil probe cooled to 30K. The use of a cryogenic probe represents an economically attractive option to enhance the sensitivity in small animal MRI/MRS.

Activity level in young patients with primary total hip arthroplasty: a 5-year minimum follow-up.

Increased activity level after total hip arthroplasty (THA) is considered a risk factor for early prosthetic failure in young patients. Forty-one primary total hip arthroplasties in 34 patients were evaluated. Walking activity was measured using a pedometer to record gait cycles. Patients completed a University of California, Los Angeles (UCLA) activity questionnaire. Linear wear rates were measured. Mean ages at surgery and final follow-up were 42 and 50.3 years, respectively (mean gait cycles per year, 1.2 million; mean UCLA score, 6; mean linear wear, 0.16 mm/y). Increased body mass index and age correlated with decreased gait cycles per year. Patients with systemic disease were less active than patients with localized hip conditions. Femoral head diameter was a predictor of linear wear. The average gait cycles per year and wear rate for this population do not appear accelerated relative to average values reported in older populations.

High-resolution capillary tube NMR. A miniaturized 5-microL high-sensitivity TXI probe for mass-limited samples, off-line LC NMR, and HT NMR.

A new triple-resonance (TXI) (1H, 13C, 15N) high-resolution nuclear magnetic resonance (NMR) capillary probe with 2.5-microL NMR-active sample volume (V(obs)) was built and tested for applications with mass- and volume-limited samples and for coupling of microbore liquid chromatography to NMR. This is the first microliter probe with optimized coil geometry for use with individual capillary tubes with an outer diameter of 1 mm. The 90 degree pulse lengths of the 1-mm microliter probe were below 2 micros for proton, below 8 micros for carbon, and below 20 micros for nitrogen, and a spectral line width at signal half-height below 1 Hz was obtained. Compared to a conventional 5-mm probe, the new 600-MHz 1-mm TXI microliter probe with z-gradient shows an increase in mass sensitivity by a factor of 5, corresponding to a 25-fold reduction in measuring time. The consumption of costly deuterated solvent is reduced by at least 2 orders of magnitude. The 1-mm TXI microliter probe with z-gradient allows the measurement of one-dimensional 1H NMR and two-dimensional heteronuclear NMR spectra with a few nanomoles (micrograms) of compound with high sensitivity, speed, and quality. This is a breakthrough for discrete sample NMR spectroscopy with paramount importance for structure elucidation in natural compound chemistry and metabolic research. It offers also advantages for linking chromatographic methods to NMR in a nindustrial environment. Capillary tube NMR may find new applications in areas where high sample throughput is essential, e.g., in the quality control of large sample arrays from parallel chemistry, screening, and compound depositories. It has the potential to increase the sample throughput by 1 order of magnitude or more if new hardware for fast sample handling and exchange becomes available.

Capsaicin-sensitive adrenal sensory fibers participate in compensatory adrenal growth in rats.

Compensatory adrenal growth, in which one gland undergoes hyperplasia after removal of the other, is mediated by a neural reflex. In the present studies, a method employing capsaicin to selectively remove adrenal sensory fibers was developed and applied to determine whether adrenal capsaicin-sensitive fibers participate in compensatory adrenal growth. The splanchnic nerves of anesthetized male rats were treated with capsaicin or vehicle. Capsaicin treatment selectively removed adrenal calcitonin gene-related peptide-positive fibers. One week after drug treatment, rats underwent left adrenalectomy or sham surgery and recovered for 5 days. Capsaicin treatment bilaterally or to the left splanchnic nerve alone (i.e., the afferent nerve in the reflex) impaired compensatory adrenal growth at 5 days compared with vehicle controls, whereas capsaicin treatment to the right splanchnic nerve alone did not affect growth. Moreover, left adrenalectomy induced c-Fos immunolabeling in ipsilateral dorsal spinal cord that was prevented by capsaicin treatment. These data suggest that adrenal capsaicin-sensitive afferent nerves participate in compensatory adrenal growth and that this effect is primarily on the afferent limb of the reflex.

Pulmonary microvascular fracture in a patient with acute respiratory distress syndrome.

OBJECTIVE: To present electron micrographs of lung tissue obtained from a patient exposed to high ventilatory pressures in the context of pulmonary dysfunction and pulmonary hypertension. DESIGN: Case report. SETTING: Adult intensive care unit of a university-affiliated teaching hospital. PATIENTS: A patient exposed to high-pressure mechanical ventilation during support for acute respiratory distress syndrome; the acute respiratory distress syndrome in this case was secondary to septic shock. MEASUREMENTS AND MAIN RESULTS: Scanning electron micrographs of lung tissue, focusing on the internal alveolar surfaces. FINDINGS: Multiple gross disruptions of the alveolar walls, suggestive of stress fractures. CONCLUSION: High-pressure mechanical ventilation may promote fracturing of the alveolar blood:airspace barrier.