GSK3β Modulates Timing-Dependent Long-Term Depression Through Direct Phosphorylation of Kv4.2 Channels.
Spike timing-dependent plasticity (STDP) is a form of activity-dependent remodeling of synaptic strength that underlies memory formation. Despite its key role in dictating learning rules in the brain circuits, the molecular mechanisms mediating STDP are still poorly understood. Here, we show that spike timing-dependent long-term depression (tLTD) and A-type K+ currents are modulated by pharmacological agents affecting the levels of active glycogen-synthase kinase 3 (GSK3) and by GSK3β knockdown in layer 2/3 of the mouse somatosensory cortex. Moreover, the blockade of A-type K+ currents mimics the effects of GSK3 up-regulation on tLTD and occludes further changes in synaptic strength. Pharmacological, immunohistochemical and biochemical experiments revealed that GSK3β influence over tLTD induction is mediated by direct phosphorylation at Ser-616 of the Kv4.2 subunit, a molecular determinant of A-type K+ currents. Collectively, these results identify the functional interaction between GSK3β and Kv4.2 channel as a novel mechanism for tLTD modulation providing exciting insight into the understanding of GSK3β role in synaptic plasticity.


Introduction of the Exotic Tomato yellow leaf curl virus-Israel in Tomato to Puerto Rico.
Thirty-five-day-old tomato plants (cultivar Florasette) exhibited yellow leaf curling, stunting, and extremely reduced fruit set in spring 2001, in Guanica, Puerto Rico (PR). Twenty percent disease incidence was observed in this field and, 8 weeks later, 75% of the plants showed symptoms. These symptoms were distinct from those caused by other tomato-infecting begomoviruses reported previously from PR, namely Merremia mosaic virus, Tomato mottle virus (ToMoV), and Potato yellow mosaic virus (1). A colony of the B biotype of Bemisia tabaci (Genn.) was used to transmit the suspect virus from symptomatic plants collected in the field and established in the greenhouse in Rio Piedras, PR. The suspect virus was transmitted readily to tomato cultivar Roma (10 of 10 plants), and symptoms were like those observed in the field. Symptoms also were reminiscent of those described for several Old World begomoviruses, referred to as Tomato yellow leaf curl virus (TYLCV). Total nucleic acids were isolated from three symptomatic field samples and three greenhouse-inoculated tomato plants showing typical disease symptoms. Extracts were analyzed by polymerase chain reaction (PCR) with primers AV2466 and AC1145 to amplify a begomoviral fragment (approximately 1.1 bp) that contains a portion of the intergenic region and the viral coat protein gene (CP) (2). Amplicons were cloned, and their nucleotide sequences were determined. A comparison of CP with other well-studied begomoviral nucleotide sequences revealed that the CP sequences for field isolates 1 to 6 shared 99.7 to 100% identity with each other and 99.9 to 100% identity with TYLCV from Israel (TYLCV-IL; accession no. X76319) as well as TYLCV-IL isolates discovered in the Dominican Republic (DO; accession no. AF024715) and, subsequently, in Florida. TYLCV-specific PCR primers (forward) 5'-GAATTCCGCCTTTAA-TTTG-3' and (reverse) 5'-GAATTCCCACTATCTTTCTC-3' were used to amplify the complete viral genome form a PR field isolate. An expected-sized amplicon of approximately 2.8 kb was obtained, and the nucleotide sequence of two cloned amplicons was determined. Genome organization revealed a predicted precoat open reading frame of 351 bp, which is characteristic of other Old World begomoviruses, including TYLCV-IL. Nucleotide comparisons indicated that the PR isolate shared 99% nucleotide sequence identity with TYLCV-IL (first reported from Israel) and introduced TYLCV-IL isolates in DO and Florida, thereby confirming the introduction of TYLCV-IL into PR. TYLCV-IL was first identified several years ago in the Western Hemisphere, and the virus has been reported in five offshore locations and three continental U.S. states since its initial introduction into the DO in the early 1990s. Considering the extreme virulence of TYLCV-IL compared with most New World tomato-infecting begomoviruses, this introduction, which likely occurred from a nearby Caribbean country or Florida, has the potential to destroy the fresh-market tomato industry in PR, which supplies tomatoes to the continental United States during the winter months. There is compelling evidence for the routine movement of tomato seedlings from the continental United States to this location in PR throughout the last 10 years, including the previous introduction of ToMoV (1). These incidences and others indicate the need for those in infected areas to take precautions to avoid further spread of this highly damaging virus in and adjacent to the Caribbean region. References: (1) A. M. Idris et al. Phytopathology 88:S42, 1998. (2) A. M. Idris and J. K. Brown, Phytopathology 88:648, 1998.


Genomic profiling of brain metastases: current knowledge and new frontiers.
Brain metastases (BM) constitute the majority of intracranial cancers and carry with them a dismal prognosis. Several common cancers have a particular predilection for spread to the brain, amongst them lung cancer, breast cancer, melanoma, renal cell carcinoma (RCC), and more rarely gastrointestinal (GI) cancers. While prognosis has historically been poor and multimodality treatment combining surgery and radiation therapy was the mainstay of treatment, the genomic revolution in cancer therapy is finding increasing applications in treatment of central nervous system (CNS) disease. Targeted therapy, combined with advances in the evaluation of BM for targetable mutations, is showing increased efficacy. Developments in the understanding of brain tropism and targetable signaling pathways in metastasis are elucidating entirely new treatment approaches. This review focuses on advances made in the understanding of the genomics of BM and how this may change the role of targeted therapeutics in this common complication of cancer.


K+-Cl- cotransporter-2 KCC2 in chicken cardiomyocytes.
Using antibodies prepared against a unique region (exon 22-24) of rat K(+)-Cl(-) cotransporter-2 (KCC2), we confirmed that the ~140-kDa KCC2 protein is exclusively expressed in rat brain, but in chicken, we observed strong reactivity not only with the ~140-kDa KCC2 protein in brain but also a slightly larger ~145-kDa protein in heart. In silico analysis showed that while exon 22 of KCC2 is unique to this isoform in therian mammals, it is retained in KCC2's closest paralog, KCC4, of lower vertebrates, including chicken. To eliminate potential cross-reactivity with chicken KCC4, the antibodies were preadsorbed with blocking peptides prepared over the only two regions showing significant sequence identity to chicken KCC4. This completely eliminated antibody recognition of exogenously expressed chicken KCC4 but not of the ~145-kDa protein in chicken heart, indicating that chicken heart expresses KCC2. Real-time PCR confirmed robust KCC2 transcript expression in both chicken brain and heart. Chicken heart expressed predominantly the longer KCC2a splice variant consistent with the larger ~145-kDa protein in chicken heart. Immunofluorescence microscopy revealed prominent plasma membrane KCC2 labeling in chicken ventricular cardiomyocytes. We hypothesize that KCC2 is an important Cl(-) extrusion pathway in avian cardiomyocytes that counters channel-mediated Cl(-) loading during high heart rates with β-adrenergic stimulation. While KCC2 is absent from mammalian cardiomyocytes, understanding the role that the other KCC isoforms play in Cl(-) homeostasis of these cells represents a nascent area of research.


Relationship between carotid artery stiffness and retinal arteriolar narrowing in healthy middle-aged persons.
Both carotid atherosclerosis and cerebral arteriolosclerosis are associated with stroke. However, the relationship between carotid atherosclerosis and cerebral arteriolosclerosis is unknown. We examined the association between carotid artery stiffness, a marker of early atherosclerosis, and retinal arteriolar narrowing, a marker of arteriolosclerosis, in healthy middle-aged people. This population-based, cross-sectional study involved 8031 men and women 45 to 64 years of age. Carotid arterial stiffness was estimated from high-resolution ultrasonic echo tracking of the left common carotid artery and was defined as adjusted arterial diameter change (AADC, micro, adjusted for diastolic blood pressure, pulse pressure and pulse pressure squared, and diastolic arterial diameter and height, with smaller AADC reflecting greater arterial stiffness). Generalized retinal arteriolar narrowing was estimated from measurements of diameters of retinal vessels from digitized retinal photographs and summarized as the arteriole-to-venule ratio (AVR, with smaller AVR indicating greater retinal arteriolar narrowing). After controlling for age, sex, ethnicity, hypertension, diabetes, and cigarette smoking, decreasing AADC was associated with decreasing AVR. The mean AADCs, comparing the lowest and highest quartiles of AVR, were 394 (SE, 4) and 409 (SE, 4) micro, respectively (P<0.01). The pattern of the graded association between carotid arterial stiffness and generalized retinal arteriolar narrowing was similar among persons with and without hypertension. Greater stiffness of the carotid arteries is related to generalized narrowing of the retinal arterioles independent of blood pressure and other vascular factors. This supports a relationship between macrovascular and microvascular disease processes important in stroke pathogenesis.


Intensive meditation training, immune cell telomerase activity, and psychological mediators.
Telomerase activity is a predictor of long-term cellular viability, which decreases with chronic psychological distress (Epel et al., 2004). Buddhist traditions claim that meditation decreases psychological distress and promotes well-being (e.g., Dalai Lama and Cutler, 2009). Therefore, we investigated the effects of a 3-month meditation retreat on telomerase activity and two major contributors to the experience of stress: Perceived Control (associated with decreased stress) and Neuroticism (associated with increased subjective distress). We used mediation models to test whether changes in Perceived Control and Neuroticism explained meditation retreat effects on telomerase activity. In addition, we investigated whether two qualities developed by meditative practice, increased Mindfulness and Purpose in Life, accounted for retreat-related changes in the two stress-related variables and in telomerase activity. Retreat participants (n=30) meditated for ∼6 h daily for 3 months and were compared with a wait-list control group (n=30) matched for age, sex, body mass index, and prior meditation experience. Retreat participants received instruction in concentrative meditation techniques and complementary practices used to cultivate benevolent states of mind (Wallace, 2006). Psychological measures were assessed pre- and post-retreat. Peripheral blood mononuclear cell samples were collected post-retreat for telomerase activity. Because there were clear, a priori hypotheses, 1-tailed significance criteria were used throughout. Telomerase activity was significantly greater in retreat participants than in controls at the end of the retreat (p<0.05). Increases in Perceived Control, decreases in Neuroticism, and increases in both Mindfulness and Purpose in Life were greater in the retreat group (p<0.01). Mediation analyses indicated that the effect of the retreat on telomerase was mediated by increased Perceived Control and decreased Neuroticism. In turn, changes in Perceived Control and Neuroticism were both partially mediated by increased Mindfulness and Purpose in Life. Additionally, increases in Purpose in Life directly mediated the telomerase group difference, whereas increases in Mindfulness did not. This is the first study to link meditation and positive psychological change with telomerase activity. Although we did not measure baseline telomerase activity, the data suggest that increases in perceived control and decreases in negative affectivity contributed to an increase in telomerase activity, with implications for telomere length and immune cell longevity. Further, Purpose in Life is influenced by meditative practice and directly affects both perceived control and negative emotionality, affecting telomerase activity directly as well as indirectly.


Dimensions of hospital nurses' quality of working life.
This paper is a report of a study describing the quality of working life of nurses in Taiwan. The purpose of the study was to gather data on which to base a questionnaire to be used in further research. Nurses often complain of overwork and underpay. Problems persist with nurses' job satisfaction, stress, organizational commitment and intent to leave. 'Quality of working life' is a system of analysing how people experience work: it relates to job satisfaction, intent to leave, turnover rate, personality and work stress. However, reliable information on hospital nurses' quality of working life is limited. A descriptive study was carried out with a convenience sample. A total of 16 focus groups in one medical centre and five regional hospitals informed a quality of working life framework. Each group had three to five participants who were Registered Nurses in medical or surgical wards with at least 2 years' nursing experience, and who held a position below assistant nurse manager. The data were collected in 2000. A total of 56 nurses' quality of working life categories were identified and fitted into six dimensions: socio-economic relevance, demography, organizational aspects, work aspects, human relation aspects and self-actualization. In this paper, we focus on issues emphasized by focus group participants. These were managing shift work within the demands of family life; accommodation; support resources; and nurses' clinical ladder system and salary system. Further research is needed with other groups of nurses in a wider variety of settings in order to examine strengths and weaknesses in the total healthcare work environment and to develop appropriate strategies for nurses' quality of working life.


Upper airway resistance syndrome, nocturnal blood pressure monitoring, and borderline hypertension.
Upper airway resistance syndrome (UARS) is a sleep-disordered breathing syndrome characterized by complaints of daytime fatigue and/or sleepiness, increased upper airway resistance during sleep, frequent transient arousals, and no significant hypoxemia. Of a population of 110 subjects (58 men) diagnosed as having UARS, we investigated acute systolic and diastolic BP changes seen during sleep in two different samples. First, six patients from the original subject pool were found to have untreated chronic borderline high BP, and were subjected to 48 h of continuous ambulatory BP monitoring before treatment and another 48 h of BP monitoring 1 month after the start of nasal-continuous positive airway pressure (N-CPAP) treatment. Five of six subjects used their equipment on a regular basis and had their chronic borderline high BP completely controlled. No change in BP values was seen in the last subject, who discontinued N-CPAP after 3 days. A second protocol investigated seven normotensive subjects drawn from the initial subject pool. Continuous radial artery BP recording was performed during nocturnal sleep with simultaneous polygraphic recording of sleep/wake variables and respiration. BP changes were studied during periods of increased respiratory efforts and at the time of alpha EEG arousals. Increases in systolic and diastolic BP were noted during the breaths with the greatest inspiratory efforts without significant hypoxemia. A further increase in BP was noted in association with arousals. Three of these subjects also underwent echocardiography during sleep, which demonstrated a leftward shift of the interventricular septum with pulsus paradoxus in association with peak end-inspiratory esophageal pressure more negative than -35 cm H2O. Our study indicates that, in the absence of classic apneas, hypopneas, and repetitive significant drops in oxygen saturation (below 90%), repetitive increases in BP can occur as a result of increased airway resistance during sleep. It also shows that, in some patients with both UARS and borderline high BP, high BP can be controlled with treatment of UARS. We conclude that abnormal upper airway resistance during sleep, often associated with snoring, can play a role in the development of hypertension.


Transfer of carbohydrates on to nascent glycoprotein of vesicular stomatitis virus.
I studied the glycosylation in vivo of a viral envelope protein, the glycoprotein of vesicular stomatitis virus (VSV), by pulse labelling of virus-infected HeLa cells with 3H-labelled monosaccharides (mannose, glucosamine). Radioactivity was incorporated into the fraction of membrane-bound polyribosomes, although metabolic conversion of [3H]-mannose into amino acids was negligible. Dissociation of bound polyribosomes revealed that the radioactively co-purified with the peptidyl-tRNA. The nascent peptides were released by alkaline hydrolysis, immunoprecipitated and analysed by polyacrylamide-gel electrophoresis. It is apparent from the size distribution of the [3H]mannose-labelled nascent chains that attachment of carbohydrate starts when approximately half of the amino acid sequence of the viral glycoprotein has been synthesized.


Prospective case-control study of serum mullerian inhibiting substance and breast cancer risk.
Müllerian inhibiting substance (MIS) is a member of the transforming growth factor beta family of growth and differentiation factors that inhibits elongation and branching of mammary ducts and has been shown to inhibit mammary tumor growth in vitro and in animal models. The objective of this study was to determine whether serum MIS levels are associated with breast cancer risk. We conducted a prospective case-control study of 309 participants who were registered in the Columbia, Missouri Serum Bank. Each of 105 in situ or invasive breast cancer case patients with prediagnostic serum collected before menopause was matched to two control subjects by age, date, menstrual cycle day, and time of day of blood collection. MIS was measured in serum by using an enzyme-linked immunosorbent assay, and estradiol and testosterone concentrations were quantified by using specific radioimmunoassays. Data were analyzed using conditional logistic regression. All tests of statistical significance were two-sided. The relative odds ratio of breast cancer for women in increasing MIS quartiles were 1, 2.8 (95% confidence interval [CI] = 1.0 to 7.4), 5.9 (95% CI = 2.4 to 14.6), and 9.8 (95% CI = 3.3 to 28.9, P(trend) < .001). The association of MIS with breast cancer was weaker in women who were not taking oral contraceptives at the time of blood collection, but adjustment for estradiol and testosterone levels did not materially alter results for these women. The association of MIS with breast cancer did not vary by age at blood collection but was stronger among women who were diagnosed with breast cancer at an older age than among those who were diagnosed at a younger age. MIS may be a novel biomarker of increased breast cancer risk. Additional research including confirmatory epidemiological studies and mechanistic studies is needed.


Mini-invasive transvaginal repair of isthmocele: a video case report.
To describe our technique for transvaginal treatment of isthmocele. Surgical video article. Local Institutional Review Board approval for the video reproduction was obtained. A scientific institute. A 26-year-old patient complaining of abnormal uterine bleeding and pelvic pain was referred to our gynecological clinic for secondary infertility. At transvaginal ultrasound examination, a cesarean scar defect of 22 × 11 mm was identified, with a residual myometrial thickness over the defect of 2 mm. Isthmocele excision and myometrial repair was performed transvaginal, under regional anesthesia. Before surgery, a hysteroscopy was performed to identify the dehiscence of the cesarean scar on the anterior wall of the uterus and to confirm the presence of the isthmocele and its distance from the external os. Then an incision was made at the anterior cervicovaginal junction and the bladder was dissected away until the anterior peritoneal reflection was identified. Hysteroscopic guidance by transillumination was used to identify the exact position and the limits of the isthmocele. The fibrotic tissue was then removed, and the myometrial defect was closed with interrupted sutures by using 2-0 Vicryl, engulfing the myometrial fibers that would tend to slide laterally. The vaginal mucosa was then sutured with interrupted Vicryl 2-0 sutures. At the end of the procedure, a hysteroscopy was performed to visualize the correction of the defect and to prove the continuity of the cervical canal with the uterine cavity. Repair of isthmocele and relief of symptoms. The postoperative course was uneventful, and the patient was discharged the day after surgery. At 1-month follow-up pelvic ultrasound showed complete anatomic repair of the uterine defect. The patient was asymptomatic with no more postmenstrual bleeding. She is satisfied with the treatment and is still trying for pregnancy. Symptomatic isthmocele can be treated surgically via a hysteroscopic, laparoscopic, or vaginal approach, depending on the clinical findings and the skill set and comfort level of the surgeon. Unfortunately, there is no consensus about the ideal surgical approach. The hysteroscopic approach has been demonstrated to be effective for the treatment of abnormal uterine bleeding; however, it does not strengthen the uterine wall and it has a risk of bladder injury. The laparoscopic approach provides good anatomic results, but it requires general anesthesia and may be associated with bladder injury. The transvaginal approach appears to be a feasible, effective, and safe modality to repair the uterine defect and to restore the original thickness of the myometrium. It is a minimally invasive, scarless, and low-cost procedure. It ensures quick recovery and a relatively pain-free postoperative course with early return to normal function.


Ant odometry in the third dimension.
Desert ants (Cataglyphis) are renowned for their ability to perform large-scale foraging excursions and then return to the nest by path integration. They do so by integrating courses steered and the distances travelled into a continually updated home vector. Whereas the angular orientation is based on skylight cues, how the ants gauge the distances travelled has remained largely unclear. Furthermore, almost all studies on path integration in Cataglyphis, as well as in spiders, rodents, and humans, have aimed at understanding how the animals compute homebound courses in the horizontal plane. Here, we investigate for the first time how an animal's odometer operates when a path integration task has to be accomplished that includes a vertical component. We trained Cataglyphis ants within arrays of uphill and downhill channels, and later tested them on flat terrain, or vice versa. In all these cases, the ants indicated homing distances that corresponded not to the distances actually travelled but to the ground distances; that is, to the sum of the horizontal projections of the uphill and downhill segments of the ants' paths.


Visual and Vestibular Inputs Affect Muscle Synergies Responsible for Body Extension and Stabilization in Sit-to-Stand Motion.
The sit-to-stand motion is a common movement in daily life and understanding the mechanism of the sit-to-stand motion is important. Our previous study shows that four muscle synergies can characterize the sit-to-stand motion, and they have specific roles, such as upper body flexion, rising from a chair, body extension, and posture stabilization. The time-varying weight of these synergies are changed to achieve adaptive movement. However, the relationship between sensory input and the activation of the muscle synergies is not completely understood. In this paper, we aim to clarify how vestibular and visual inputs affect the muscle synergy in sit-to-stand motion. To address this, we conducted experiments as follows. Muscle activity, body kinematics, and ground reaction force were measured for the sit-to-stand motion under three different conditions: control, visual-disturbance, and vestibular-disturbance conditions. Under the control condition, the participants stood without any intervention. Under the visual-disturbance condition, the participants wore convex lens glasses and performed the sit-to-stand motion in a dark room. Under the vestibular-disturbance condition, a caloric test was performed. Muscle synergies were calculated for these three conditions using non-negative matrix factorization. We examined whether the same four muscle synergies were employed under each condition, and the changes in the time-varying coefficients were determined. These experiments were conducted on seven healthy, young participants. It was found that four muscle synergies could explain the muscle activity in the sit-to-stand motion under the three conditions. However, there were significant differences in the time-varying weight coefficients. When the visual input was disturbed, a larger amplitude was found for the muscle synergy that activated mostly in the final posture stabilization phase of the sit-to-stand motion. Under vestibular-disturbance condition, a longer activation was observed for the synergies that extended the entire body and led to posture stabilization. The results implied that during human sit-to-stand motion, visual input has less contribution to alter or correct activation of muscle synergies until the last phase. On the other hand, duration of muscle synergies after the buttocks leave are prolonged in order to adapt to the unstable condition in which sense of verticality is decreased under vestibular-disturbance.


Polycomb repressor complex-2 is a novel target for mesothelioma therapy.
Polycomb group (PcG) proteins are critical epigenetic mediators of stem cell pluripotency, which have been implicated in the pathogenesis of human cancers. This study was undertaken to examine the frequency and clinical relevance of PcG protein expression in malignant pleural mesotheliomas (MPM). Microarray, quantitative reverse transcriptase PCR (qRT-PCR), immunoblot, and immunohistochemistry techniques were used to examine PcG protein expression in cultured MPM, mesothelioma specimens, and normal mesothelial cells. Lentiviral short hairpin RNA techniques were used to inhibit EZH2 and EED expression in MPM cells. Proliferation, migration, clonogenicity, and tumorigenicity of MPM cells either exhibiting knockdown of EZH2 or EED, or exposed to 3-deazaneplanocin A (DZNep), and respective controls were assessed by cell count, scratch and soft agar assays, and murine xenograft experiments. Microarray and qRT-PCR techniques were used to examine gene expression profiles mediated by knockdown of EZH2 or EED, or DZNep. EZH2 and EED, which encode components of polycomb repressor complex-2 (PRC-2), were overexpressed in MPM lines relative to normal mesothelial cells. EZH2 was overexpressed in approximately 85% of MPMs compared with normal pleura, correlating with diminished patient survival. Overexpression of EZH2 coincided with decreased levels of miR-101 and miR-26a. Knockdown of EZH2 orEED, or DZNep treatment, decreased global H3K27Me3 levels, and significantly inhibited proliferation, migration, clonogenicity, and tumorigenicity of MPM cells. Common as well as differential gene expression profiles were observed following knockdown of PRC-2 members or DZNep treatment. Pharmacologic inhibition of PRC-2 expression/activity is a novel strategy for mesothelioma therapy.


Structural Basis of the Substrate Selectivity of Viperin.
Viperin is a radical S-adenosylmethionine (SAM) enzyme that inhibits viral replication by converting cytidine triphosphate (CTP) into 3'-deoxy-3',4'-didehydro-CTP and by additional undefined mechanisms operating through its N- and C-terminal domains. Here, we describe crystal structures of viperin bound to a SAM analogue and CTP or uridine triphosphate (UTP) and report kinetic parameters for viperin-catalyzed reactions with CTP or UTP as substrates. Viperin orients the C4' hydrogen atom of CTP and UTP similarly for abstraction by a 5'-deoxyadenosyl radical, but the uracil moiety introduces unfavorable interactions that prevent tight binding of UTP. Consistently, kcat is similar for CTP and UTP whereas the Km for UTP is much greater. The structures also show that nucleotide binding results in ordering of the C-terminal tail and reveal that this region contains a P-loop that binds the γ-phosphate of the bound nucleotide. Collectively, the results explain the selectivity for CTP and reveal a structural role for the C-terminal tail in binding CTP and UTP.


Inapplicability of fractional moving blood volume technique to standardize Virtual Organ Computer-aided AnaLysis indices for quantified three-dimensional power Doppler.
To determine whether the technique of fractional moving blood volume (FMBV) is applicable to Virtual Organ Computer-aided AnaLysis II (VOCAL II™)-based indices to quantify three-dimensional power Doppler ultrasound (3D-PDU) by investigating the effect of gain level on the indices measured at a possible reference point for standardization. Ten women with singleton pregnancy between 33+3 and 37+5 weeks' gestation were recruited. The optimal position for 3D acquisition of cord insertion into the placenta was identified and static 3D-PDU volumes were acquired using consistent machine configurations. Without moving the probe or the participant changing position, successive 3D volumes were stored at -3, -5, -7 and -9 dB and at the individualized sub-noise gain (SNG) level. Volumes were excluded if flash artifact was present, in which case all five volumes were reacquired. Using 4D View software, the cord insertion was magnified and the smallest sphere possible was used to measure vascularization index (VI), flow index (FI) and vascularization flow index (VFI). The associations between VOCAL indices and gain level were assessed using Pearson's correlation coefficient. VOCAL indices for cord insertion correlated poorly with gain level, whether fundamental or relative to SNG level (R(2) = 0.07 and 0.04, respectively). VI was consistently 100% and mean FI and VFI were 99.5 (SD, 0.57), with all values > 97 irrespective of gain level. Whilst previous work has shown that gain correlates well with placental tissue VOCAL indices, the correlation between gain level and VOCAL indices in an area of 100% vascularity at the cord insertion is poor. Regions of 100% vascularity appear to be artificially assigned a value approaching 100% for all VOCAL indices irrespective of gain level. This precludes using the technique of VOCAL indices from large vessels to standardize power Doppler measurements and the FMBV index is therefore not applicable to image analysis using VOCAL.


Efficient iPS cell production with the MyoD transactivation domain in serum-free culture.
A major difficulty of producing induced pluripotent stem cells (iPSCs) has been the low efficiency of reprogramming differentiated cells into pluripotent cells. We previously showed that 5% of mouse embryonic fibroblasts (MEFs) were reprogrammed into iPSCs when they were transduced with a fusion gene composed of Oct4 and the transactivation domain of MyoD (called M(3)O), along with Sox2, Klf4 and c-Myc (SKM). In addition, M(3)O facilitated chromatin remodeling of pluripotency genes in the majority of transduced MEFs, including cells that did not become iPSCs. These observations suggested the possibility that more than 5% of cells had acquired the ability to become iPSCs given more favorable culture conditions. Here, we raised the efficiency of making mouse iPSCs with M(3)O-SKM to 26% by culturing transduced cells at low density in serum-free culture medium. In contrast, the efficiency increased from 0.1% to only 2% with the combination of wild-type Oct4 and SKM (OSKM) under the same culture condition. For human iPSCs, M(3)O-SKM achieved 7% efficiency under a similar serum-free culture condition, in comparison to 1% efficiency with OSKM. This study highlights the power of combining the transactivation domain of MyoD with a favorable culture environment.


d-Penicillamine-coated Cu/Ag alloy nanocluster superstructures: aggregation-induced emission and tunable photoluminescence from red to orange.
The aggregation-induced emission (AIE) behavior of metal nanoclusters has attracted much attention owing to their extensive application prospects in bio-imaging and chemical sensors. However, the intrinsic mechanism of metal nanoclusters' aggregation-induced emission is still not very clear. Herein, Cu nanoclusters S1 and Ag(i)-doped Cu/Ag nanoclusters S2 and S3 coated with d-penicillamine are designed and synthesized by a self-assembly strategy. S1-S3 show strong luminescence properties with luminescence quantum yields as high as 11.4%-14.2%. Moreover, their luminescence peak position shows an obvious hypsochromic shift from 615 (S1) to 570 nm (S3). With the introduction of Ag(i) ions, the assembly morphology also shows clear changes from the irregular assembly structure (S1) to large spherical particles with the average size of 0.18 μm for S2 and 0.47 μm for S3. A detailed investigation of high-resolution transmission electron microscopy (HRTEM) images, X-ray diffraction (XRD) patterns, ESI-TOF-mass spectra, UV-vis absorption spectra, photoluminescence spectra and luminescence lifetimes of Cu/Ag nanoclusters S2 and S3 indicates that Cu/Ag nanoclusters are actually a Cu/Ag alloy nanocluster superstructure and the microscopic arrangement of S2 and S3 is more compact and ordered relative to S1. The more compact and ordered Cu/Ag alloy nanocluster superstructure enhances the metal-metal interaction of inter-nanoclusters and intra-nanoclusters and facilitates the radiative transition of ligand-to-metal charge transfer (LMCT) and/or ligand-to-metal-metal charge transfer (LMMCT), which results in the aggregation-induced emission phenomenon. In addition, an enhanced metal-metal interaction increases the average metal-metal distance of the Cu/Ag nanocluster superstructure, leading to a hypsochromic shift of emission spectra. Furthermore, the Cu/Ag alloy nanoclusters show good stability and reversibility on pH cycling between pH = 3 and 7 and temperature cycling between 5 °C and 48 °C. The Cu/Ag alloy nanoclusters can be used as probes for Ag(i) ion and halide anion detection in real water samples by the ratiometric PL method (I570/I615).


What happens before? A field experiment exploring how pay and representation differentially shape bias on the pathway into organizations.
Little is known about how discrimination manifests before individuals formally apply to organizations or how it varies within and between organizations. We address this knowledge gap through an audit study in academia of over 6,500 professors at top U.S. universities drawn from 89 disciplines and 259 institutions. In our experiment, professors were contacted by fictional prospective students seeking to discuss research opportunities prior to applying to a doctoral program. Names of students were randomly assigned to signal gender and race (White, Black, Hispanic, Indian, Chinese), but messages were otherwise identical. We hypothesized that discrimination would appear at the informal "pathway" preceding entry to academia and would vary by discipline and university as a function of faculty representation and pay. We found that when considering requests from prospective students seeking mentoring in the future, faculty were significantly more responsive to White males than to all other categories of students, collectively, particularly in higher-paying disciplines and private institutions. Counterintuitively, the representation of women and minorities and discrimination were uncorrelated, a finding that suggests greater representation cannot be assumed to reduce discrimination. This research highlights the importance of studying decisions made before formal entry points into organizations and reveals that discrimination is not evenly distributed within and between organizations.


Trabecular bone turnover, bone marrow cell development, and gene expression of bone matrix proteins after low calcium feeding in rats.
Low-calcium-fed animals have been accepted as one of the experimental models showing a reduction in bone mass. However, the effects of short-term low-calcium feeding on bone turnover, the development of osteoprogenitor cells, and gene expression of bone matrix proteins have not been reported. In this study, we examined the effect of a low-calcium diet on rat tibia and analyzed the changes in the bone by histomorphometry, bone marrow cell culture, and in situ and Northern hybridization of the bone matrix proteins. Rats were fed either a low-calcium diet (0.05% Ca) or a normal calcium diet (0.5% Ca) using the pair feeding technique. They were killed at day 0, 12 h, and days 1, 2, and 3. In the low-calcium group, the serum parathyroid hormone (PTH) level was temporarily increased in 12 h after feeding the low-calcium diet. Bone mineral density in the trabecular bone was significantly decreased from 1 day after the low-calcium feeding, but cortical bone did not show any changes during the experimental period. The bone volume per tissue volume in the proximal tibia also decreased from day 1 in the low-calcium group. The number of osteoclasts and osteoblasts on the trabecular bone surface was increased in the low-calcium group compared with the normal-calcium group. An ex vivo study showed that the number of progenitors of osteoclasts and osteoblasts in bone marrow was also increased in the low-calcium group of rats. The localization of type I collagen mRNA was observed in osteoblasts in the low-calcium group. The Northern hybridization study showed that the gene expression of type I collagen, osteopontin, and osteocalcin was increased at day 3 in the low-calcium group. These results indicated that the trabecular bone surface quickly responded to the low-calcium feeding and that bone remodeling activity was activated probably by PTH. The changes in bone marrow cell populations and the gene expression of bone matrix proteins are closely associated with increased bone turnover induced by the low-calcium diet, resulting in rapid bone loss of the trabecular bone.


A composite biochemical system for bacterial nitrate and nitrite assimilation as exemplified by Paracoccus denitrificans.
The denitrifying bacterium Paracoccus denitrificans can grow aerobically or anaerobically using nitrate or nitrite as the sole nitrogen source. The biochemical pathway responsible is expressed from a gene cluster comprising a nitrate/nitrite transporter (NasA), nitrite transporter (NasH), nitrite reductase (NasB), ferredoxin (NasG) and nitrate reductase (NasC). NasB and NasG are essential for growth with nitrate or nitrite as the nitrogen source. NADH serves as the electron donor for nitrate and nitrite reduction, but only NasB has a NADH-oxidizing domain. Nitrate and nitrite reductase activities show the same Km for NADH and can be separated by anion-exchange chromatography, but only fractions containing NasB retain the ability to oxidize NADH. This implies that NasG mediates electron flux from the NADH-oxidizing site in NasB to the sites of nitrate and nitrite reduction in NasC and NasB respectively. Delivery of extracellular nitrate to NasBGC is mediated by NasA, but both NasA and NasH contribute to nitrite uptake. The roles of NasA and NasC can be substituted during anaerobic growth by the biochemically distinct membrane-bound respiratory nitrate reductase (Nar), demonstrating functional overlap. nasG is highly conserved in nitrate/nitrite assimilation gene clusters, which is consistent with a key role for the NasG ferredoxin, as part of a phylogenetically widespread composite nitrate and nitrite reductase system.


Patient indicators of a pulmonary exacerbation: preliminary reports from school aged children map onto those of adults.
Despite the importance of identifying and managing a pulmonary exacerbation, and its use as an outcome measure in interventions, there is no standardised definition in cystic fibrosis. In achieving standardised criteria it is important to identify patient-reported indicators. Interviews were undertaken with 35 school aged children. They reported symptoms experienced during a pulmonary exacerbation in two ways: the first symptoms they become aware of, and how they recognised when they were improving. Interviews were taped, transcribed verbatim and the data analysed thematically. For many children, the onset of an exacerbation was characterised by 'cold' symptoms, tiredness, and changes in cough. For those with moderate or severe disease, sleep disruption, activity induced breathlessness, changes in mood, sputum volume and lack of appetite were common. When describing improvement children focused initially on activities they were now able to perform accompanied by improvements in tiredness and cough. Those with moderate or severe disease also reported improvements in sleep and mood, breathlessness, sputum volume and colour. Child-reported indicators of a pulmonary exacerbation tend to map onto those reported by adults. These results provide the rationale for the development of a single scale for school age children and adults that could be sensitive to progressive stages of CF disease.


Low intensity microwave radiation induced oxidative stress, inflammatory response and DNA damage in rat brain.
Over the past decade people have been constantly exposed to microwave radiation mainly from wireless communication devices used in day to day life. Therefore, the concerns over potential adverse effects of microwave radiation on human health are increasing. Until now no study has been proposed to investigate the underlying causes of genotoxic effects induced by low intensity microwave exposure. Thus, the present study was undertaken to determine the influence of low intensity microwave radiation on oxidative stress, inflammatory response and DNA damage in rat brain. The study was carried out on 24 male Fischer 344 rats, randomly divided into four groups (n=6 in each group): group I consisted of sham exposed (control) rats, group II-IV consisted of rats exposed to microwave radiation at frequencies 900, 1800 and 2450 MHz, specific absorption rates (SARs) 0.59, 0.58 and 0.66 mW/kg, respectively in gigahertz transverse electromagnetic (GTEM) cell for 60 days (2h/day, 5 days/week). Rats were sacrificed and decapitated to isolate hippocampus at the end of the exposure duration. Low intensity microwave exposure resulted in a frequency dependent significant increase in oxidative stress markers viz. malondialdehyde (MDA), protein carbonyl (PCO) and catalase (CAT) in microwave exposed groups in comparison to sham exposed group (p<0.05). Whereas, levels of reduced glutathione (GSH) and superoxide dismutase (SOD) were found significantly decreased in microwave exposed groups (p<0.05). A significant increase in levels of pro-inflammatory cytokines (IL-2, IL-6, TNF-α, and IFN-γ) was observed in microwave exposed animal (p<0.05). Furthermore, significant DNA damage was also observed in microwave exposed groups as compared to their corresponding values in sham exposed group (p<0.05). In conclusion, the present study suggests that low intensity microwave radiation induces oxidative stress, inflammatory response and DNA damage in brain by exerting a frequency dependent effect. The study also indicates that increased oxidative stress and inflammatory response might be the factors involved in DNA damage following low intensity microwave exposure.


Physician-reported readiness to change stroke prevention practices.
There are a series of possible impediments to the incorporation of new treatment modalities into clinical practice, and any intervention intended to alter practice must consider physicians' motivation and readiness to change. As part of a national survey in the United States, physicians from a variety of specialties were asked whether they were comfortable with, considering changing or expecting to make changes in their screening and treatment practices for a series of eight hypothetical patients at elevated risk of stroke. Readiness to change varied with the type of patient under consideration and with physician specialty, but not with a series of other physician and practice characteristics. Knowledge of physicians' states of readiness to change in combination with data relating to current practices and potential barriers to implementation should aid in targeting educational efforts and in the development of specific interventions to improve stroke prevention.


Variable Expressivity in Fragile X Syndrome: Towards the Identification of Molecular Characteristics That Modify the Phenotype.
Fragile X syndrome (FXS), is an X-linked inherited genetic disease. FXS is the leading cause of inherited intellectual disability and autism in the world. Those affected are characterized by intellectual disability, language deficit, typical facies, and macroorchidism. Alterations in the FMR1 gene have been associated with FXS. The majority of people with this condition have an allele with an expansion of more than 200 repeats in a tract of CGGs within the 5' untranslated region, and this expansion is associated with a hypermethylated state of the gene promoter. FXS has incomplete penetrance and variable expressivity. Intellectual disability is present in 100% of males and 60% of females. Autism spectrum disorder symptoms appear in 50% to 60% of males and 20% of females. Other characteristics such as behavioral and physical alterations have significant variations in presentation frequency. The molecular causes of the variable phenotype in FXS patients are becoming clear: these causes are related to the FMR1 gene itself and to secondary, modifying gene effects. In FXS patients, size and methylation mosaicisms are common. Secondary to mosaicism, there is a variation in the quantity of FMR1 mRNA and the protein coded by the gene Fragile Mental Retardation Protein (FMRP). Potential modifier genes have also been proposed, with conflicting results. Characterizing patients according to CGG expansion, methylation status, concentration of mRNA and FMRP, and genotypification for possible modifier genes in a clinical setting offers an opportunity to identify predictors for treatment response evaluation. When intervention strategies become available to modulate the course of the disease they could be crucial for selecting patients and identifying the best therapeutic intervention. The purpose of this review is to present the information available about the molecular causes of the variability of the expression incomplete penetrance and variable expressivity in FXS and their potential clinical applications.


Global meta-analysis shows pervasive phosphorus limitation of aboveground plant production in natural terrestrial ecosystems.
Phosphorus (P) limitation of aboveground plant production is usually assumed to occur in tropical regions but rarely elsewhere. Here we report that such P limitation is more widespread and much stronger than previously estimated. In our global meta-analysis, almost half (46.2%) of 652 P-addition field experiments reveal a significant P limitation on aboveground plant production. Globally, P additions increase aboveground plant production by 34.9% in natural terrestrial ecosystems, which is 7.0-15.9% higher than previously suggested. In croplands, by contrast, P additions increase aboveground plant production by only 13.9%, probably because of historical fertilizations. The magnitude of P limitation also differs among climate zones and regions, and is driven by climate, ecosystem properties, and fertilization regimes. In addition to confirming that P limitation is widespread in tropical regions, our study demonstrates that P limitation often occurs in other regions. This suggests that previous studies have underestimated the importance of altered P supply on aboveground plant production in natural terrestrial ecosystems.


Exploring obstacles to critical care trials in the UK: A qualitative investigation.
Clinical trials in critical care are often resource-intense, with many unique challenges. Barriers to effective recruitment and implementation of study intervention have not been explored in a UK context. To identify facilitating factors and barriers to enrolling patients into critical care clinical trials within the UK from clinician's perspectives. A qualitative interview study was undertaken on behalf of the National Institute of Health Research critical care specialty group, in which research active clinicians across different Clinical Research Networks were interviewed. A loosely structured interview schedule was used, based on themes generated from the literature associated with accessing critical care trials. Research teams (critical care doctors, research nurses, and trial coordinators) from hospitals from each Clinical Research Network were contacted to try to achieve representation across the UK. Interviews were carried out across nine UK Clinical Research Networks with a range of doctors and research nurses. All hospitals were teaching hospitals with varying research nurse numbers and allocated consultant research sessions. There were a range of six to nine ongoing clinical trials in critical care for each centre representative interviewed. Data were analysed using framework analysis, and six final themes were identified related to factors associated with: centre, unit, resources, study, clinician, and patient/family. The most commonly cited barrier to conducting clinical trials was related to resources, namely insufficient human and financial resources, leading to staff and study recruitment difficulties. Clinical uncertainty and equipoise regarding comparative merits of trials were challenging in terms of engaging critical care teams. A number of patient and family factors added complexities in terms of recruitment; however, refusal rates were generally reported as low. Flexibility in funding and employment by research teams enables continuity of studies and staff. Innovative measures to incentivise research nurses and clinical teams can help recruit more patients into trials. Research teams are highly committed to providing cover to recruit critical care trials, and a significant effort to anticipate barriers is undertaken; these endeavours are summarised to provide guidance for other teams wishing to address any potential difficulties.


The functional neuroanatomy of decision-making.
Decision-making is a complex executive function that draws on past experience, present goals, and anticipation of outcome, and which is influenced by prevailing and predicted emotional tone and cultural context. Functional imaging investigations and focal lesion studies identify the orbitofrontal, anterior cingulate, and dorsolateral prefrontal cortices as critical to decision-making. The authors review the connections of these prefrontal regions with the neocortex, limbic system, basal ganglia, and cerebellum, highlight current ideas regarding the cognitive processes of decision-making that these networks subserve, and present a novel integrated neuroanatomical model for decision-making. Finally, clinical relevance of this circuitry is illustrated through a discussion of frontotemporal dementia, traumatic brain injury, and sociopathy.


Risk factors and outcomes for recurrent paediatric in-hospital cardiac arrest: Retrospective multicenter cohort study.
Recurrent in-hospital cardiac arrest (IHCA) is associated with morbidity and mortality in adults. We aimed to describe the risk factors and outcomes for paediatric recurrent IHCA. Retrospective cohort study of patients ≤18 years old with single or recurrent IHCA. Recurrent IHCA was defined as ≥2 IHCA within the same hospitalization. Categorical variables expressed as percentages and compared via Chi square test. Continuous variables expressed as medians with interquartile ranges and compared via rank sum test. Outcomes assessed in a propensity match cohort. From July 1, 2015 to January 26, 2021, 139/894 (15.5%) patients experienced recurrent IHCA. Compared to patients with a single IHCA, recurrent IHCA patients were more likely to be trauma and less likely to be surgical cardiac patients. Median duration of cardiopulmonary resuscitation (CPR) was shorter in the recurrent IHCA (5 vs. 11 min; p < 0.001) with no difference in IHCA location or immediate cause of CPR. Patients with recurrent IHCA had worse survival to intensive care unit (ICU) discharge (31% vs. 52%; p < 0.001), and worse survival to hospital discharge (30% vs. 48%; p < 0.001) in unadjusted analyses and after propensity matching, patients with recurrent IHCA still had worse survival to ICU (34% vs. 67%; p < 0.001) and hospital (31% vs. 64%; p < 0.001) discharge. When examining those with a single vs. a recurrent IHCA, event and patient factors including more pre-existing conditions and shorter duration of CPR were associated with risk for recurrent IHCA. Recurrent IHCA is associated with worse survival outcomes following propensity matching.


Liver disease does not affect lipolysis as measured with the 13C-mixed triacylglycerol breath test in children with cystic fibrosis.
Liver disease associated with cystic fibrosis may not only limit the solubilisation and absorption of the products of fat digestion, but also may depress the activity of pancreatic lipase. The purpose of this study was to measure the effect of liver disease on triacylglycerol lipolysis using the 13C-mixed triacylglycerol breath test. Forty children with cystic fibrosis took 13C-mixed triacylglycerol with a standard breakfast and the child's normal pancreatic enzyme replacement therapy. Breath samples were collected before and every 30 minutes after ingestion for 6 hours. The cumulative percentage dose of 13C recovered at 6 hours was calculated from sequential measurements of 13C enrichment of breath CO2, measured by isotope ratio mass spectrometry. Liver abnormalities and portal hypertension were defined by ultrasound scan and clinical examination. Twenty-four children had liver abnormalities, including 5 with portal hypertension. No difference was found between cumulative percentage dose of 13C recovered at 6 hours in 16 children with no liver abnormality (mean, 21.4%+/-11.1%), 19 children with liver abnormalities (22.2%+/-10.0%) and 5 children with portal hypertension (20.9%+/-7.1%). Intestinal lipolysis is not reduced in cystic fibrosis liver disease when measured using the 13C mixed triacylglycerol breath test. These findings affirm the test's use as an indirect measure of fat digestion that is not affected by inadequate intraluminal bile salts or liver disease.


Risk posed to honeybees (Apis mellifera L, Hymenoptera) by an imidacloprid seed dressing of sunflowers.
In a greenhouse metabolism study, sunflowers were seed-treated with radiolabelled imidacloprid in a 700 g kg-1 WS formulation (Gaucho WS 70) at 0.7 mg AI per seed, and the nature of the resulting residues in nectar and pollen was determined. Only the parent compound and no metabolites were detected in nectar and pollen of these seed-treated sunflower plants (limit of detection < 0.001 mg kg-1). In standard LD50 laboratory tests, imidacloprid showed high oral toxicity to honeybees (Apis mellifera), with LD50 values between 3.7 and 40.9 ng per bee, corresponding to a lethal food concentration between 0.14 and 1.57 mg kg-1. The residue level of imidacloprid in nectar and pollen of seed-treated sunflower plants in the field was negligible. Under field-growing conditions no residues were detected (limit of detection: 0.0015 mg kg-1) in either nectar or pollen. There were also no detectable residues in nectar and pollen of sunflowers planted as a succeeding crop in soils which previously had been cropped with imidacloprid seed-treated plants. Chronic feeding experiments with sunflower honey fortified with 0.002, 0.005, 0.010 and 0.020 mg kg-1 imidacloprid were conducted to assess potential long-term adverse effects on honeybee colonies. Testing end-points in this 39-day feeding study were mortality, feeding activity, wax/comb production, breeding performance and colony vitality. Even at the highest test concentration, imidacloprid showed no adverse effects on the development of the exposed bee colonies. This no-adverse-effect concentration of 0.020 mg kg-1 compares with a field residue level of less than 0.0015 mg kg-1 (= limit of detection in the field residue studies) which clearly shows that a sunflower seed dressing with imidacloprid poses no risk to honeybees. This conclusion is confirmed by observations made in more than 10 field studies and several tunnel tests.


Multiple dimensions of dietary diversity in large mammalian herbivores.
Theory predicts that trophic specialization (i.e. low dietary diversity) should make consumer populations sensitive to environmental disturbances. Yet diagnosing specialization is complicated both by the difficulty of precisely quantifying diet composition and by definitional ambiguity: what makes a diet 'diverse'? We sought to characterize the relationship between taxonomic dietary diversity (TDD) and phylogenetic dietary diversity (PDD) in a species-rich community of large mammalian herbivores in a semi-arid East African savanna. We hypothesized that TDD and PDD would be positively correlated within and among species, because taxonomically diverse diets are likely to include plants from many lineages. By using DNA metabarcoding to analyse 1,281 faecal samples collected across multiple seasons, we compiled high-resolution diet profiles for 25 sympatric large-herbivore species. For each of these populations, we calculated TDD and PDD with reference to a DNA reference library for local plants. Contrary to our hypothesis, measures of TDD and PDD were either uncorrelated or negatively correlated with each other. Thus, these metrics reflect distinct dimensions of dietary specialization both within and among species. In general, grazers and ruminants exhibited greater TDD, but lower PDD, than did browsers and non-ruminants. We found significant seasonal variation in TDD and/or PDD for all but four species (Grevy's zebra, buffalo, elephant, Grant's gazelle); however, the relationship between TDD and PDD was consistent across seasons for all but one of the 12 best-sampled species (plains zebra). Our results show that taxonomic generalists can be phylogenetic specialists, and vice versa. These two dimensions of dietary diversity suggest contrasting implications for efforts to predict how consumers will respond to climate change and other environmental perturbations. For example, populations with low TDD may be sensitive to phylogenetically 'random' losses of food species, whereas populations with low PDD may be comparatively more sensitive to environmental changes that disadvantage entire plant lineages-and populations with low dietary diversity in both taxonomic and phylogenetic dimensions may be most vulnerable of all.


A method for detecting single mRNA molecules in Arabidopsis thaliana.
Despite advances in other model organisms, there are currently no techniques to explore cell-to-cell variation and sub-cellular localization of RNA molecules at the single-cell level in plants. Here we describe a method for imaging individual mRNA molecules in Arabidopsis thaliana root cells using multiple singly labeled oligonucleotide probes. We demonstrate detection of both mRNA and nascent transcripts of the housekeeping gene Protein Phosphatase 2A. Our image analysis pipeline also enables quantification of mRNAs that reveals the frequency distribution of transcripts per cell underlying the population mean. This method allows single molecule RNA in situ to be exploited as a powerful tool for studying gene regulation in plants.


Comparing Oncological Outcomes and Surgical Complications of Hand-Assisted, Laparoscopic and Robotic Nephroureterectomy for Upper Tract Urothelial Carcinoma.
This study aimed to compare the oncological outcomes and surgical complications of patients with upper tract urothelial carcinoma (UTUC) treated with different minimally invasive techniques for nephroureterectomy. From the updated data of the Taiwan UTUC Collaboration Group, a total of 3,333 UTUC patients were identified. After excluding ineligible cases, we retrospectively included 1,340 patients from 15 institutions who received hand-assisted laparoscopic nephroureterectomy (HALNU), laparoscopic nephroureterectomy (LNU) or robotic nephroureterectomy (RNU) between 2001 and 2021. Kaplan-Meier estimator and Cox proportional hazards model were used to analyze the survival outcomes, and binary logistic regression model was selected to compare the risks of postoperative complications of different surgical approaches. Among the enrolled patients, 741, 458 and 141 patients received HALNU, LNU and RNU, respectively. Compared with RNU (41.1%) and LNU (32.5%), the rate of lymph node dissection in HALNU was the lowest (17.4%). In both Kaplan-Meier and univariate analysis, the type of surgery was significantly associated with overall and cancer-specific survival. The statistical significance of surgical methods on survival outcomes remained in multivariate analysis, where patients undergoing HALNU appeared to have the worst overall (p = 0.007) and cancer-specific (p = 0.047) survival rates among the three groups. In all analyses, the surgical approach was not related to bladder recurrence. In addition, HALNU was significantly associated with longer hospital stay (p = 0.002), and had the highest risk of major Clavien-Dindo complications (p = 0.011), paralytic ileus (p = 0.012), and postoperative end-stage renal disease (p <0.001). Minimally invasive surgery can be safe and feasible. We proved that compared with the HALNU group, the LNU and RNU groups have better survival rates and fewer surgical complications. It is crucial to uphold strict oncological principles with sophisticated technique to improve outcomes. Further prospective studies are needed to validate our findings.


The public health critical race methodology: praxis for antiracism research.
The number of studies targeting racial health inequities and the capabilities for measuring racism effects have grown substantially in recent years. Still, the need remains for a public health framework that moves beyond merely documenting disparities toward eliminating them. Critical Race Theory (CRT) has been the dominant influence on racial scholarship since the 1980s; however, its jurisprudential origins have, until now, limited its application to public health research. To improve the ease and fidelity with which health equity research applies CRT, this paper introduces the Public Health Critical Race praxis (PHCR). PHCR aids the study of contemporary racial phenomena, illuminates disciplinary conventions that may inadvertently reinforce social hierarchies and offers tools for racial equity approaches to knowledge production.


Cloning and nucleotide sequence of the thrB gene from the cyanobacterium Calothrix PCC 7601.
The cyanobacterium Calothrix PCC 7601 thrB gene, encoding homoserine kinase (EC 18.104.22.168), was cloned via complementation of an Escherichia coli threonine auxotroph, and its nucleotide sequence was determined. The comparison of the homoserine kinase amino acid sequences from Calothrix PCC 7601, E. coli K12 and Bacillus subtilis 168 indicates a closer relationship between cyanobacteria and bacillaceae than between cyanobacteria and enterobacteriaceae. Sequence analysis of the 5' and 3' flanking regions of the Calothrix thrB gene revealed the existence of a 169-codon-long open reading frame downstream from thrB: this sequence may be the second gene of a Calothrix thr operon. Two types of tandemly repeated sequences, sharing similarities with other prokaryotic transcriptional regulatory elements, were detected in the region upstream from the thrB gene.


Presenilin-1 affects trafficking and processing of betaAPP and is targeted in a complex with nicastrin to the plasma membrane.
Amyloid beta-peptide (Abeta) is generated by the consecutive cleavages of beta- and gamma-secretase. The intramembraneous gamma-secretase cleavage critically depends on the activity of presenilins (PS1 and PS2). Although there is evidence that PSs are aspartyl proteases with gamma-secretase activity, it remains controversial whether their subcellular localization overlaps with the cellular sites of Abeta production. We now demonstrate that biologically active GFP-tagged PS1 as well as endogenous PS1 are targeted to the plasma membrane (PM) of living cells. On the way to the PM, PS1 binds to nicastrin (Nct), an essential component of the gamma-secretase complex. This complex is targeted through the secretory pathway where PS1-bound Nct becomes endoglycosidase H resistant. Moreover, surface-biotinylated Nct can be coimmunoprecipitated with PS1 antibodies, demonstrating that this complex is located to cellular sites with gamma-secretase activity. Inactivating PS1 or PS2 function by mutagenesis of one of the critical aspartate residues or by gamma-secretase inhibitors results in delayed reinternalization of the beta-amyloid precursor protein and its accumulation at the cell surface. Our data suggest that PS is targeted as a biologically active complex with Nct through the secretory pathway to the cell surface and suggest a dual function of PS in gamma-secretase processing and in trafficking.


Zinc uptake and HMA4 activity are required for micro- and macroelement balance in tobacco (Nicotiana tabacum).
The pleiotropic effects of zinc deficiency on ion homeostasis have already been described in several plants. Tobacco (Nicotiana tabacum) heavy metal ATPases HMA4.1 and HMA4.2 are involved in zinc and cadmium root-to-shoot translocation. In previous research, we have shown that N. tabacum HMA4 RNAi plants and HMA4 double-nonsense mutants exhibit strongly reduced zinc and cadmium levels in leaves as well as stunted growth. In this study, the ionome and transcriptome of these lines were investigated to better characterize the effect of reduced zinc levels and to understand the impaired growth phenotype. We found that, under standard greenhouse fertilization rates, these lines accumulated up to 4- to 6-fold more phosphorus, iron, manganese, and copper than their respective controls. Under field conditions, HMA4 double-mutant plants also exhibited similar accumulation phenotypes, albeit to a lower extent. In both HMA4 RNAi plants and HMA4 mutants, transcription analysis showed a local zinc-deficiency response in leaves as well as an FIT1-mediated iron-deficiency response in roots, likely contributing to iron and manganese uptake at the root level. A phosphate-starvation response involving HHO2 was also observed in HMA4-impaired plant leaves. The high level of phosphorus observed in HMA4-impaired plants is correlated with leaf swelling and necrosis. The upregulation of aquaporin genes is in line with cellular water influx and the observed leaf swelling phenotype. These results highlight the involvement of HMA4 in zinc homeostasis and related regulatory processes that balance the micro- and macroelements in above-ground organs.


Pathological hypertrophy and cardiac interstitium. Fibrosis and renin-angiotensin-aldosterone system.
Left ventricular hypertrophy (LVH) is the major risk factor associated with myocardial failure. An explanation for why a presumptive adaptation such as LVH would prove pathological has been elusive. Insights into the impairment in contractility of the hypertrophied myocardium have been sought in the biochemistry of cardiac myocyte contraction. Equally compelling is a consideration of abnormalities in myocardial structure that impair organ contractile function while preserving myocyte contractility. For example, in the LVH that accompanies hypertension, the extracellular space is frequently the site of an abnormal accumulation of fibrillar collagen. This reactive and progressive interstitial and perivascular fibrosis accounts for abnormal myocardial stiffness and ultimately ventricular dysfunction and is likely a result of cardiac fibroblast growth and enhanced collagen synthesis. The disproportionate involvement of this nonmyocyte cell, however, is not a uniform accompaniment to myocyte hypertrophy and LVH, suggesting that the growth of myocyte and nonmyocyte cells is independent of each other. This has now been demonstrated in in vivo studies of experimental hypertension in which the abnormal fibrous tissue response was found in the hypertensive, hypertrophied left ventricle as well as in the normotensive, nonhypertrophied right ventricle. These findings further suggest that a circulating substance that gained access to the common coronary circulation of the ventricles was involved. This hypothesis has been tested in various animal models in which plasma concentrations of angiotensin II and aldosterone were varied. Based on morphometric and morphological findings, it can be concluded that arterial hypertension (i.e., an elevation in coronary perfusion pressure) together with elevated circulating aldosterone are associated with cardiac fibroblast involvement and the resultant heterogeneity in tissue structure. Nonmyocyte cells of the cardiac interstitium represent an important determinant of pathological LVH. The mechanisms that invoke short- (e.g., collagen metabolism) and long-term (e.g., mitosis) responses of cardiac fibroblasts require further investigation and integration of in vitro with in vivo studies. The stage is set, however, to prevent pathological LVH resulting from myocardial fibrosis as well as to reverse it.


Measuring surgical outcomes in congenital craniofacial surgery: an objective approach.
Assessment of surgical outcome in congenital craniofacial malformations is necessary to evaluate treatment efficiency since the results of early surgery are influenced by surgical technique and timing. Available outcome measurements all have their limitations, especially for application in congenital craniofacial malformations. Therefore a new outcome measurement was developed, in which each facial unit is scored in a standardized way. For each facial unit, deformities of shape or contour, malposition and soft tissue involvement were evaluated, besides scoring for specific congenital malformations of that area. The final result was tested on pre- and postoperative photographs of patients with rare facial clefts and reliability and validity were demonstrated. The new developed instrument showed the ability to provide quantification of outcome. Because of its properties it can serve as an instrument to compare outcome between techniques, surgeons and centers in a more objective and standardized way.


Bio-templated fabrication of metal-free boron carbonitride tubes for visible light photocatalysis.
The synthesis of boron carbon nitride (BCN) with special morphologies via a simple pathway is still a major challenge. A novel and facile biotemplating method has been presented to synthesize boron carbon nitride tubes (BCNTs) by using kapok fibers (KFs) as both biotemplates and carbonaceous sources. The BCNT photocatalysts can catalyse hydrogen evolution from water under visible light illumination.


Blood will out: vascular contributions to Alzheimer's disease.
The fundamental pathology in Alzheimer's disease (AD) is neuronal dysfunction leading to cognitive impairment. The amyloid-β peptide (Aβ), derived from amyloid precursor protein, is one driver of AD, but how it leads to neuronal dysfunction is not established. In this Review, I discuss the complexity of AD and possible cause-and-effect relationships between Aβ and the vascular and hemostatic systems. AD can be considered a multifactorial syndrome with various contributing pathological mechanisms. Therefore, as is routinely done with cancer, it will be important to classify patients with respect to their disease signature so that specific pathologies, including vascular pathways, can be therapeutically targeted.


Parents' Readiness to Change Affects BMI Reduction Outcomes in Adolescents with Polycystic Ovary Syndrome.
Evidence supports the importance of parental involvement for youth's ability to manage weight. This study utilized the stages of change (SOC) model to assess readiness to change weight control behaviors as well as the predictive value of SOC in determining BMI outcomes in forty adolescent-parent dyads (mean adolescent age = 15 ± 1.84 (13-20), BMI = 37 ± 8.60; 70% white) participating in a weight management intervention for adolescent females with polycystic ovary syndrome (PCOS). Adolescents and parents completed a questionnaire assessing their SOC for the following four weight control domains: increasing dietary portion control, increasing fruit and vegetable consumption, decreasing dietary fat, and increasing usual physical activity. Linear regression analyses indicated that adolescent change in total SOC from baseline to treatment completion was not predictive of adolescent change in BMI from baseline to treatment completion. However, parent change in total SOC from baseline to treatment completion was predictive of adolescent change in BMI, (t(24) = 2.15, p = 0.043). Findings support future research which carefully assesses adolescent and parent SOC and potentially develops interventions targeting adolescent and parental readiness to adopt healthy lifestyle goals.


Human plague: An old scourge that needs new answers.
Yersinia pestis, the bacterial causative agent of plague, remains an important threat to human health. Plague is a rodent-borne disease that has historically shown an outstanding ability to colonize and persist across different species, habitats, and environments while provoking sporadic cases, outbreaks, and deadly global epidemics among humans. Between September and November 2017, an outbreak of urban pneumonic plague was declared in Madagascar, which refocused the attention of the scientific community on this ancient human scourge. Given recent trends and plague's resilience to control in the wild, its high fatality rate in humans without early treatment, and its capacity to disrupt social and healthcare systems, human plague should be considered as a neglected threat. A workshop was held in Paris in July 2018 to review current knowledge about plague and to identify the scientific research priorities to eradicate plague as a human threat. It was concluded that an urgent commitment is needed to develop and fund a strong research agenda aiming to fill the current knowledge gaps structured around 4 main axes: (i) an improved understanding of the ecological interactions among the reservoir, vector, pathogen, and environment; (ii) human and societal responses; (iii) improved diagnostic tools and case management; and (iv) vaccine development. These axes should be cross-cutting, translational, and focused on delivering context-specific strategies. Results of this research should feed a global control and prevention strategy within a "One Health" approach.


Injuries among Swedish female elite football players: a prospective population study.
Injury occurrence in all 12 female elite senior football teams in premiere league was registered during 1 year. Of 269 players, 129 (48%) sustained 237 injuries. The total injury incidence was 4.6/1000 h of football. The injury incidence during practice was 2.7 and during the game time was 13.9/1000 h. The highest injury incidence during practice was to the knee (0.8/1000 h) and thigh (0.5/1000 h), and during game time was to the knee (4.4/1000 h) and head (2.2/1000 h). In total, the location for the highest injury incidence was the knee with 1.5 injuries/1000 h of football. The majority of injuries (82%) were localized to the lower extremities. Sixty-six injuries (28%) were re-occurring injuries (re-injuries). The incidence of traumatic injuries was 3.3/1000 h of football and for overuse injuries 1.3/1000 h. Overuse injuries occurred mainly during the preseason and at the beginning of the spring season. Thirty-nine percent of the injuries were slight or minor causing absence from practice or game time of less than 1 week, 39% were moderate (absence 7-28 days) and 22% were major (absence more than 28 days). The major injuries occurred often owing to trauma and were mainly to the knee.


A chaperone cascade sorts proteins for posttranslational membrane insertion into the endoplasmic reticulum.
Tail-anchored (TA) proteins are posttranslationally inserted into either the endoplasmic reticulum (ER) or the mitochondrial outer membrane. The C-terminal transmembrane domains (TMDs) of TA proteins enable their many essential cellular functions by specifying the membrane target, but how cells process these targeting signals is poorly understood. Here, we reveal the composition of a conserved multiprotein TMD recognition complex (TRC) and show that distinct TRC subunits recognize the two types of TMD signals. By engineering mutations in a mitochondrial TMD, we switch over its TRC subunit recognition, thus leading to its misinsertion into the ER. Biochemical reconstitution with purified components demonstrates that TRC tethers and enzymatically activates Get3 to selectively hand off ER-bound TA proteins to Get3. Thus, ER-bound TA proteins are sorted at the top of a TMD chaperone cascade that culminates with the formation of Get3-TA protein complexes, which are recruited to the ER membrane for insertion.


Voluntarily reported prescribing, monitoring and medication transfer errors in intensive care units in The Netherlands.
Background Medication errors occur frequently in intensive care units (ICU). Voluntarily reported medication errors form an easily available source of information. Objective This study aimed to characterize prescribing, monitoring and medication transfer errors that were voluntarily reported in the ICU, in order to reveal medication safety issues. Setting This retrospective data analysis study included reports of medication errors from eleven Dutch ICU's from January 2016 to December 2017. Method We used data extractions from the incident reporting systems of the participating ICU's. The reports were transferred into one database and categorized into type of error, cause, medication (groups), and patient harm. Descriptive statistics were used to calculate the proportion of medication errors and the distribution of subcategories. Based on the analysis, ICU medication safety issues were revealed. Main outcome measure The main outcome measure was the proportion of prescribing, monitoring and medication transfer error reports. Results Prescribing errors were reported most frequently (n = 233, 33%), followed by medication transfer errors (n = 85, 12%) and monitoring errors (n = 27, 4%). Other findings were: medication transfer errors frequently caused serious harm, especially the omission of home medication involving the central nervous system and proton pump inhibitors; omissions and dosing errors occurred most frequently; protocol problems caused a quarter of the medication errors; and medications needing blood level monitoring (e.g. tacrolimus, vancomycin, heparin and insulin) were frequently involved. Conclusion This analysis of voluntarily reported prescribing, monitoring and medication transfer errors warrants several improvement measures in these processes, which may help to increase medication safety in the ICU.


Mx1 but not MxA confers resistance against tick-borne Dhori virus in mice.
The interferon-induced nuclear Mx1 protein is responsible for innate resistance of mice to influenza virus. It has been unclear why mice are equipped with a powerful and specific defense mechanism against influenza viruses for which they are not natural hosts. Here, we show that Dhori virus, an influenza-like virus transmitted by ticks and known to infect small mammals, is sensitive to the Mx1 resistance mechanism. Influenza virus-susceptible BALB/c and C57BL/6 mice (lacking a functional Mx1 gene) developed severe disease symptoms and died within a few days after intraperitoneal infection with a lethal dose of Dhori virus. In contrast, Mx1(+)-congenic, influenza virus-resistant BALB.A2G-Mx1 and B6.A2G-Mx1 mice remained healthy and survived. The Mx1 resistance phenotype was expressed in cultured peritoneal macrophages and interferon-treated embryonic cells obtained from these mice. Moreover, stable lines of transfected mouse 3T3 cells constitutively expressing Mx1 protein were protected from Dhori virus infection. The MxA protein of human cells shows a high degree of sequence similarity to Mx1 but, unlike Mx1, inhibits a broad range of RNA viruses. Transgenic mice that permanently express the human MxA protein in various organs became resistant to infection with Thogoto virus but remained fully susceptible to Dhori virus. These in vivo results show that DHO virus is unique in being resistant to human MxA but susceptible to mouse Mx1 protein. They further indicate that the Mx1 system functions as a potent defense mechanism against tick-borne influenza-like viruses in mice.


Experimental and theoretical studies on aqueous-phase reactivity of hydroxyl radicals with multiple carboxylated and hydroxylated benzene compounds.
In this study, we shed light on the initial addition of hydroxyl radicals (HO˙) to multiple carboxylated and hydroxylated benzene compounds in aqueous-phase advanced oxidation processes (AOPs). We analyze the experimentally measured transient spectra near neutral pH using quantum mechanical-based time-dependent density functional theory (TD-DFT). The ab initio DFT method was first used to find and optimize aqueous-phase transition state structures, then the TD-DFT was used to analyze molecular orbitals (MOs) of the optimized transition state structures to reveal the functional groups that are responsible for the individual absorption peaks. The initial addition of HO˙ to the benzene ring produced hydroxycyclohexadienyl radicals. Then, HO-adducts are generated from dimerization or disproportionation of hydroxycyclohexadienyl radicals and represent their transient spectral peaks at approximately 350 nm and 250 nm. As reaction proceeds, the HO-adducts are decreased depending on the subsequent reactions. These investigations into the experimental transient spectra coupled with the theoretical analysis using the TD-DFT enable us to visualize an initial transformation of organic compounds induced by the aqueous phase HO˙ oxidation. Moreover, the experimental reaction rate constants and the theoretically calculated aqueous phase free energies of activation provide quantitative insights into the addition of HO˙ to multiple carboxylated and hydroxylated benzene compounds.


Deep learning assessment of breast terminal duct lobular unit involution: Towards automated prediction of breast cancer risk.
Terminal duct lobular unit (TDLU) involution is the regression of milk-producing structures in the breast. Women with less TDLU involution are more likely to develop breast cancer. A major bottleneck in studying TDLU involution in large cohort studies is the need for labor-intensive manual assessment of TDLUs. We developed a computational pathology solution to automatically capture TDLU involution measures. Whole slide images (WSIs) of benign breast biopsies were obtained from the Nurses' Health Study. A set of 92 WSIs was annotated for acini, TDLUs and adipose tissue to train deep convolutional neural network (CNN) models for detection of acini, and segmentation of TDLUs and adipose tissue. These networks were integrated into a single computational method to capture TDLU involution measures including number of TDLUs per tissue area, median TDLU span and median number of acini per TDLU. We validated our method on 40 additional WSIs by comparing with manually acquired measures. Our CNN models detected acini with an F1 score of 0.73±0.07, and segmented TDLUs and adipose tissue with Dice scores of 0.84±0.13 and 0.87±0.04, respectively. The inter-observer ICC scores for manual assessments on 40 WSIs of number of TDLUs per tissue area, median TDLU span, and median acini count per TDLU were 0.71, 0.81 and 0.73, respectively. Intra-observer reliability was evaluated on 10/40 WSIs with ICC scores of >0.8. Inter-observer ICC scores between automated results and the mean of the two observers were: 0.80 for number of TDLUs per tissue area, 0.57 for median TDLU span, and 0.80 for median acini count per TDLU. TDLU involution measures evaluated by manual and automated assessment were inversely associated with age and menopausal status. We developed a computational pathology method to measure TDLU involution. This technology eliminates the labor-intensiveness and subjectivity of manual TDLU assessment, and can be applied to future breast cancer risk studies.


Nanometric axial resolution of fibronectin assembly units achieved with an efficient reconstruction approach for multi-angle-TIRF microscopy.
High resolution imaging of molecules at the cell-substrate interface is required for understanding key biological processes. Here we propose a complete pipeline for multi-angle total internal reflection fluorescence microscopy (MA-TIRF) going from instrument design and calibration procedures to numerical reconstruction. Our custom setup is endowed with a homogeneous field illumination and precise excitation beam angle. Given a set of MA-TIRF acquisitions, we deploy an efficient joint deconvolution/reconstruction algorithm based on a variational formulation of the inverse problem. This algorithm offers the possibility of using various regularizations and can run on graphics processing unit (GPU) for rapid reconstruction. Moreover, it can be easily used with other MA-TIRF devices and we provide it as an open-source software. This ensemble has enabled us to visualize and measure with unprecedented nanometric resolution, the depth of molecular components of the fibronectin assembly machinery at the basal surface of endothelial cells.


Mutations in DNA polymerase gamma cause error prone DNA synthesis in human mitochondrial disorders.
This paper summarizes recent advances in understanding the links between the cell's ability to maintain integrity of its mitochondrial genome and mitochondrial genetic diseases. Human mitochondrial DNA is replicated by the two-subunit DNA polymerase gamma (polgamma). We investigated the fidelity of DNA replication by polgamma with and without exonucleolytic proofreading and its p55 accessory subunit. Polgamma has high base substitution fidelity due to efficient base selection and exonucleolytic proofreading, but low frameshift fidelity when copying homopolymeric sequences longer than four nucleotides. Progressive external ophthalmoplegia (PEO) is a rare disease characterized by the accumulation of large deletions in mitochondrial DNA. Recently, several mutations in the polymerase and exonuclease domains of the human polgamma have been shown to be associated with PEO. We are analyzing the effect of these mutations on the human polgamma enzyme. In particular, three autosomal dominant mutations alter amino acids located within polymerase motif B of polgamma. These residues are highly conserved among family A DNA polymerases, which include T7 DNA polymerase and E.coli pol I. These PEO mutations have been generated in polgamma to analyze their effects on overall polymerase function as well as the effects on the fidelity of DNA synthesis. One mutation in particular, Y955C, was found in several families throughout Europe, including one Belgian family and five unrelated Italian families. The Y955C mutant polgamma retains a wild-type catalytic rate but suffers a 45-fold decrease in apparent binding affinity for the incoming dNTP. The Y955C derivative is also much less accurate than is wild-type polgamma, with error rates for certain mismatches elevated by 10- to 100-fold. The error prone DNA synthesis observed for the Y955C polgamma is consistent with the accumulation of mtDNA mutations in patients with PEO. The effects of other polgamma mutations associated with PEO are discussed.


Evaluation of the safety and efficacy of periodontal applications of a living tissue-engineered human fibroblast-derived dermal substitute. II. Comparison to the subepithelial connective tissue graft: a randomized controlled feasibility study.
The subepithelial connective tissue graft, traditionally harvested from the patient's palate, is commonly used for root coverage in periodontal recession defects. This study evaluates the safety and effectiveness of a living human fibroblast-derived dermal substitute (HF-DDS) compared to a connective tissue graft (CTG) for root coverage in these situations. Thirteen patients were selected for this study. Each patient had Miller Class I or II bilateral facial recession defects > or =3 mm on two non-adjacent teeth. The test tooth received an HF-DDS graft, while a CTG was placed on the control site. The 10 test surgeries were performed by one operator and three pilot surgeries were performed by another surgeon. Eight of the HF-DDS sites received a single thickness of material; five received a double thickness. Clinical measurements were taken at baseline; 1 week; and 1, 3, and 6 months following surgery. Parameters measured were plaque index, recession depth, clinical attachment levels, recession width, probing depth, and width of keratinized tissue. All clinical readings were taken by a masked, calibrated examiner. There were no statistically significant differences between the test and control groups. The amount of root coverage was slightly greater for the control group than for the test group, but statistically the difference was insignificant. The width of the recession defect measured at the cemento-enamel junction (CEJ) for the test group was slightly smaller than that of the control group at the conclusion of the study. The amount of keratinized tissue was the same in both groups at 6 months. The probing depth was slightly greater in the control group as was the gain in clinical attachment, but neither was statistically significant. The amount of root coverage obtained when one layer of HF-DDS was used compared to the amount of coverage obtained when two layers were used approached statistical significance, but the small sample size may have been responsible for the difference. Within the limits of this study, the human fibroblast-derived dermal substitute may offer potential as a substitute to the connective tissue graft for covering areas of facial Miller Class I or Class II gingival recession in humans.


Super-mini percutaneous nephrolithotomy (PCNL) vs standard PCNL for the management of renal calculi of <2 cm: a randomised controlled study.
To compare the effectiveness and safety of standard percutaneous nephrolithotomy (sPCNL) and super-mini PCNL (SMP). A total of 150 patients presenting with renal calculi of <2 cm were randomised to either sPCNL (Group 1) or SMP (Group 2). Randomisation was based on centralised computer-generated numbers. Variables studied included: stone-free rates (SFRs), operative time, intra- and postoperative complications, postoperative pain score, analgesic requirement, and hospital stay. Statistical analysis was performed using a t-test or Mann-Whitney U-test for continuous variables and chi-squared test or Fisher's exact test for categorical variables. Between September 2018 and April 2019, 75 patients were included in each group. The SFRs of the groups were similar (97.33 vs 98.66%, P = 0.56). The mean (sd) operative time was significantly longer in Group 2, at 36.40 (14.07) vs 23.12 (11.96) min (P < 0.001). The mean (sd) decrease in haemoglobin was significantly less in Group 2, at 3.0 (4.9) vs 7.5 (6.5) g/L (P < 0.001). The mean (sd) pain score at 24 h was significantly lower in Group 2, at 0.3 (0.46) vs 0.75 (0.53) (P < 0.001). The mean (sd) analgesic requirement was significantly less in Group 2, at 67 (22.49) vs 91.5 (30.56) mg tramadol (P < 0.001). The mean (sd) hospital stay was significantly less in Group 2, at 28.38 (3.6) vs 39.84 (3.7) h (P < 0.001). SMP is equally as effective as sPCNL for managing renal calculi of <2 cm, with improved safety. Although SMP is associated with a longer operative time, it has a significantly lower incidence of bleeding and postoperative pain, and a shorter hospital stay.


Training for health care in developing countries: the work of the Tropical Health and Education Trust.
The Tropical Health and Education Trust (THET) was established to strengthen medical education and training for health care in developing countries. The Trust responds to requests from training institutions with a wide range of activities and programmes. Projects to meet specific needs are planned in outline with the Deans or Directors of institutions, as a basis for a long-term link with a similar institution in the United Kingdom. These links are now the preferred method for meeting requests to develop skills, strengthen services and promote staff development. However, funding is always necessary for their support. THET has promoted students' community-based training by enabling students in a team-training programme in Ethiopia to make interventions in primary health care. A prize for the best students' community, clinical or laboratory projects in six African countries encourages enquiry by the students, promotes independent learning, and relates academic work to problems in health care. Work with Ministries of Health includes a continuing medical education programme for rural medical officers in Uganda, courses in basic and life-saving surgery for Ethiopian health and medical officers, and a programme to update the skills of laboratory technologists in rural hospitals in Ghana. The range of projects that THET supports is wide because the needs, defined by those who are working in, and responsible for, training in the health service are diverse.


Conformational Dynamics of Mechanically Compliant DNA Nanostructures from Coarse-Grained Molecular Dynamics Simulations.
Structural DNA nanotechnology, the assembly of rigid 3D structures of complex yet precise geometries, has recently been used to design dynamic, mechanically compliant nanostructures with tunable equilibrium conformations and conformational distributions. Here we use coarse-grained molecular dynamics simulations to provide insights into the conformational dynamics of a set of mechanically compliant DNA nanostructures-DNA hinges that use single-stranded DNA "springs" to tune the equilibrium conformation of a layered double-stranded DNA "joint" connecting two stiff "arms" constructed from DNA helix bundles. The simulations reproduce the experimentally measured equilibrium angles between hinge arms for a range of hinge designs. The hinges are found to be structurally stable, except for some fraying of the open ends of the DNA helices comprising the hinge arms and some loss of base-pairing interactions in the joint regions coinciding with the crossover junctions, especially in hinges designed to exhibit a small bending angle that exhibit large local stresses resulting in strong kinks in their joints. Principal component analysis reveals that while the hinge dynamics are dominated by bending motion, some twisting and sliding of hinge arms relative to each other also exists. Forced deformation of the hinges reveals distinct bending mechanisms for hinges with short, inextensible springs versus those with longer, more extensible springs. Lastly, we introduce an approach for rapidly predicting equilibrium hinge angles from individual force-deformation behaviors of its single- and double-stranded DNA components. Taken together, these results demonstrate that coarse-grained modeling is a promising approach for designing, predicting, and studying the dynamics of compliant DNA nanostructures, where conformational fluctuations become important, multiple deformation mechanisms exist, and continuum approaches may not yield accurate properties.


A Peptide-Based Mechano-sensitive, Proteolytically Stable Hydrogel with Remarkable Antibacterial Properties.
A long-chain amino acid containing dipeptide has been found to form a hydrogel in phosphate buffer whose pH ranges from 6.0 to 8.8. The hydrogel formed at pH 7.46 has been characterized by small-angle X-ray scattering (SAXS), wide-angle powder X-ray diffraction (PXRD), Fourier transform infrared (FT-IR) spectroscopy, field-emission scanning electron microscopy (FE-SEM), high-resolution transmission electron microscopy (HR-TEM) imaging and rheological analyses. The microscopic imaging studies suggest the formation of a nanofibrillar three-dimensional (3D) network for the hydrogel. As observed visually and confirmed rheologically, the hydrogel at pH 7.46 exhibits thixotropy. This thixotropic property can be exploited to inject the peptide. Furthermore, the hydrogel exhibits remarkable antibacterial activity against Gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa, which are responsible for many common diseases. The hydrogel has practical applicability due to its biocompatibility with human red blood cells and human fibroblast cells. Interestingly, this hydrogel shows high resistance toward proteolytic enzymes, making it a new potential antimicrobial agent for future applications. It has also been observed that a small change in molecular structure of the gelator peptide not only turns the gelator into a nongelator molecule under similar conditions, but it also has a significant negative impact on its bactericidal character.