Oxytocin/neurophysin-I messenger ribonucleic acid in bovine granulosa cells increases after the luteinizing hormone (LH) surge and is stimulated by LH in vitro.

Bovine granulosa cells express the oxytocin/neurophysin-I (OT/NP-I) gene and secrete OT in vitro. We have shown previously that bovine granulosa cells isolated from the preovulatory follicle after the LH surge secrete 20 times more OT over 5 days in culture than granulosa cells obtained before the surge. LH or FSH stimulates OT secretion in vitro by granulosa cells isolated before the LH surge. We also observed that granulosa cells of preovulatory follicles isolated before the LH surge respond to OT with an increase in progesterone secretion, suggesting that OT may be involved in regulating the follicular/luteal phase shift, or ovulation, in an autocrine fashion. The objective of this study was to determine whether the increase in OT secretion from granulosa cells after the LH surge is regulated at the level of mRNA accumulation, peptide synthesis, and/or peptide secretion. Bovine preovulatory follicles were obtained during the early follicular phase (approximately 36 h before the LH surge), during the midfollicular phase (approximately 12 h before the LH surge), or during the late follicular phase (after the LH surge). Total RNA isolated from granulosa cells and theca interna at the time of cell isolation or after culture with or without LH was subjected to Northern analysis for OT/NP-I mRNA and quantified by densitometry. OT/NP-I mRNA was not detectable or was barely detectable in granulosa cells collected during the early or midfollicular phase (n = 6 and n = 4 follicles, respectively), but a strong hybridization signal was obtained from RNA isolated after the LH surge (n = 5 follicles; P < 0.01). In contrast, OT/NP-I mRNA was not detectable in theca interna before or after the LH surge. Although OT/NP-I mRNA was not detectable in granulosa cells isolated 24 h after prostaglandin F2 alpha injection, after 24 h in culture, a weak OT/NP-I mRNA hybridization signal was observed in RNA from granulosa cells in LH-containing cultures. After 72 h in culture, granulosa cells cultured in control, as well as in LH-containing medium, exhibited a strong signal for OT/NP-I mRNA, but granulosa cells treated with LH exhibited a stronger OT/NP-I hybridization signal than control cultures (P < 0.01). Theca interna did not yield any OT/NP-I hybridization signal initially, and none was induced in culture.(ABSTRACT TRUNCATED AT 400 WORDS)     

Systemic sclerosis: patients' perceptions of their condition

OBJECTIVE: To examine patients' beliefs about systemic sclerosis (SSc) and to investigate the relationship between these beliefs, symptom report, and clinical and demographic variables. METHODS: A total of 49 patients (7 male, 42 female) with SSc underwent clinical examination and completed the Revised Illness Perception Questionnaire. This measure assesses beliefs about symptoms, chronicity or recurrence of the condition, consequences, personal and treatment control, illness coherence, perceived causes of the condition, and patients' emotional response to their condition. RESULTS: The symptoms patients most frequently associated with their SSc were stiff joints (79%), pain (75%), and fatigue (75%). The most commonly reported causes of SSc were stress (53%), altered immunity (49%), and chance or bad luck (46%). More than 96% of patients believed that their condition would be chronic and 78% believed that the condition had serious consequences on their lives. Patients with diffuse cutaneous SSc reported more significant consequences of the condition and less personal control of their SSc compared with patients with limited cutaneous disease. There were no significant differences in illness beliefs between patients with nonsevere and severe ischemia. Multiple regression analyses indicated that illness beliefs, in particular perceived consequences associated with the condition, accounted for a significant proportion of the variance in emotional response to the condition. CONCLUSIONS: The beliefs held and symptoms experienced by patients with SSc are not ruled by disease subtype, skin score, functional ability, or severity of digital ischemia. This suggests patients' beliefs and emotional response are associated with the meaning they ascribe to their condition rather than its severity.     

Rv3615c is a highly immunodominant RD1 (Region of Difference 1)-dependent secreted antigen specific for Mycobacterium tuberculosis infection

The 6-kDa early secretory antigenic target of Mycobacterium tuberculosis (ESAT-6) and the 10-kDa culture filtrate antigen (CFP-10), encoded in region of difference 1 (RD1) and secreted by the ESAT-6 system 1 (Esx-1) secretion system, are the most immunodominant and highly M. tuberculosis (MTB)-specific antigens. These attributes are responsible for their primary importance in tuberculosis (TB) immunodiagnosis and vaccine development. Rv3615c [Esx-1 substrate protein C (EspC)], encoded outside RD1, is similar in size and sequence homology to CFP-10 and ESAT-6, suggesting it might be a target of cellular immunity in TB. Using ex vivo enzyme-linked immunospot- and flow cytometry-based cytokine-secretion assay, we comprehensively assessed cellular immune responses to EspC in patients with active TB, latently infected persons, and uninfected bacillus Calmette-Guérin (BCG)-vaccinated controls. EspC was at least as immunodominant as ESAT-6 and CFP-10 in both active and latent TB infection. EspC contained broadly recognized CD4(+) and CD8(+) epitopes, inducing a predominantly CD4(+) T-cell response that comprised functional T-cell subsets secreting both IFN-γ and IL-2 as well as functional T-cell subsets secreting only IFN-γ. Surprisingly, T-cell responses to EspC were as highly specific (93%) for MTB infection as responses to ESAT-6 and CFP-10, with only 2 of 27 BCG-vaccinated controls responding to each antigen. Using quantitative proteomics and metabolically labeled mutant and genetically complemented MTB strains, we identified the mechanism of the specificity of anti-EspC immunity as the Esx-1 dependence of EspC secretion. The high immunodominance of EspC, equivalent to that of ESAT-6 and CFP-10, makes it a TB vaccine candidate, and its high specificity confers strong potential for T-cell-based immunodiagnosis.     

Original research: Predictors and outcomes of acute respiratory failure in hospitalised patients with acute pancreatitis

Background and aimAcute pancreatitis (AP) is associated with organ failures and systemic complications, most commonly acute respiratory failure (ARF) and acute kidney injury. So far, no studies have analysed the predictors and hospitalisation outcomes, of patients with AP who developed ARF. The aim of this study was to measure the prevalence of ARF in AP and to determine the clinical predictors for ARF and mortality in AP.MethodsThis is a retrospective cohort study using the Nationwide Inpatient Sample database from the year 2005–2014. The study population consisted of all hospitalisations with a primary or secondary discharge diagnosis of AP, which is further stratified based on the presence of ARF. The outcome measures include in-hospital mortality, hospital length of stay and hospitalisation cost.ResultsIn our study, about 5.4% of patients with AP had a codiagnosis of ARF, with a mortality rate of 26.5%. The significant predictors for ARF include sepsis, pleural effusion, pneumonia and cardiogenic shock. Key variables that were associated with a higher risk of mortality include mechanical ventilation, age more than 65 years, sepsis and cancer (excluding pancreatic cancer). The presence of ARF increased hospital stay by 8.3 days and hospitalisation charges by US$103 460.ConclusionIn this study, we demonstrate that ARF is a significant risk factor for increased hospital mortality, greater length of stay and higher hospitalisation charges in patients with AP. This underlines significantly higher resource utilisation in patients with a dual diagnosis of AP-ARF.     

From the Cover: Neurophysiological coordination of duet singing

Coordination of behavior for cooperative performances often relies on linkages mediated by sensory cues exchanged between participants. How neurophysiological responses to sensory information affect motor programs to coordinate behavior between individuals is not known. We investigated how plain-tailed wrens (Pheugopedius euophrys) use acoustic feedback to coordinate extraordinary duet performances in which females and males rapidly take turns singing. We made simultaneous neurophysiological recordings in a song control area “HVC” in pairs of singing wrens at a field site in Ecuador. HVC is a premotor area that integrates auditory feedback and is necessary for song production. We found that spiking activity of HVC neurons in each sex increased for production of its own syllables. In contrast, hearing sensory feedback produced by the bird’s partner decreased HVC activity during duet singing, potentially coordinating HVC premotor activity in each bird through inhibition. When birds sang alone, HVC neurons in females but not males were inhibited by hearing the partner bird. When birds were anesthetized with urethane, which antagonizes GABAergic (γ-aminobutyric acid) transmission, HVC neurons were excited rather than inhibited, suggesting a role for GABA in the coordination of duet singing. These data suggest that HVC integrates information across partners during duets and that rapid turn taking may be mediated, in part, by inhibition.

Animals routinely rely on sensory feedback for the control of their own behavior. In cooperative performances, such sensory feedback can include cues produced by other participants (1–8). For example, in interactive vocal communication, including human speech, individuals take turns vocalizing. This “turn taking” is a consequence of each participant responding to auditory cues from a partner (4–6, 9, 10). The role of such “heterogenous” (other-generated) feedback in the control of vocal turn taking and other cooperative performances is largely unknown.Plain-tailed wrens (Pheugopedius euophrys) are neotropical songbirds that cooperate to produce extraordinary duet performances but also sing by themselves (Fig. 1A) (4, 10, 11). Singing in plain-tailed wrens is performed by both females and males and used for territorial defense and other functions, including mate guarding and attraction (1, 11–16). During duets, female and male plain-tailed wrens take turns, alternating syllables at a rate of between 2 and 5 Hz (Fig. 1A) (4, 11).Open in a separate windowFig. 1.Neural control of solo and duet singing in plain-tailed wrens. (A) Spectrogram of a singing bout that included male solo syllables (blue line, top) followed by a duet. Solo syllables for both sexes (only male solo syllables are shown here) are sung at lower amplitudes than syllables produced in duets. Note that the smeared appearance of wren syllables in spectrograms reflects the acoustic structure of plain-tailed wren singing. (B and C) Each bird has a motor system that is used to produce song and sensory systems that mediate feedback. (B) During solo singing, the bird hears its own song, which is known as autogenous feedback (orange). (C) During duet singing, each bird hears both its own singing and the singing of its partner, known as heterogenous feedback (green). The key difference between solo and duet singing is heterogenous feedback that couples the neural systems of the two birds. This coupling results in changes in syllable amplitude and timing in both birds.There is a categorical difference between solo and duet singing. In solo singing, the singing bird receives only autogenous (hearing its own vocalization) feedback (Fig. 1B). The partner may hear the solo song if it is nearby, a heterogenous (other-generated) cue. In duet singing, birds receive both heterogenous and autogenous feedback as they alternate syllable production (Fig. 1C). Participants use heterogenous feedback during duet singing for precise timing of syllable production (4, 11). For example, when a male temporarily stops participating in a duet, the duration of intersyllable intervals between female syllables increases (4), showing an effect of heterogenous feedback on the timing of syllable production.How does the brain of each wren integrate heterogenous acoustic cues to coordinate the precise timing of syllable production between individuals during duet performances? To address this question, we examined neurophysiological activity in HVC, a nucleus in the nidopallium [an analogue of mammalian cortex (17, 18)]. HVC is necessary for song learning, production, and timing in species of songbirds that do not perform duets (19–24). Neurons in HVC are active during singing and respond to playback of the bird’s own learned song (25–27). In addition, recent work has shown that HVC is also involved in vocal turn taking (19).To examine the role of heterogenous feedback in the control of duet performances, we compared neurophysiological activity in HVC when female or male wrens sang solo syllables with syllables sung during duets. Neurophysiological recordings were made in awake and anesthetized pairs of wrens at the Yanayacu Biological Station and Center for Creative Studies on the slopes of the Antisana volcano in Ecuador. We found that heterogenous cues inhibited HVC activity during duet performances in both females and males, but inhibition was only observed in females during solo singing.     

Gonadotropin-induced expression of messenger ribonucleic acid for cyclooxygenase-2 and production of prostaglandins E and F2alpha in bovine preovulatory follicles are regulated by the progesterone receptor

Follicular production of prostaglandins (PGs) is essential for ovulation, but the factors mediating gonadotropin-induced secretion of PGE and PGF(2alpha) remain largely unknown. We tested the hypothesis that gonadotropin-induced changes in progesterone and its receptor (PR) mediate the increase in periovulatory PGs. Heifers were treated with PGF(2alpha) and GnRH to induce luteolysis and the LH/FSH surge (ovulation occurs approximately 30 h after GnRH). Because there are two increases in intrafollicular progesterone/PR mRNA during the bovine periovulatory period, we first examined the temporal pattern of PG production by follicles collected at 0, 3.5, 6, 12, 18, and 24 h after GnRH. Although PGs did not increase in the follicular fluid until 24 h after GnRH, acute secretion of PGs by follicle wall (theca + granulosa cells) was initiated by 18 h and had increased manyfold by 24 h after GnRH. In vitro, FSH and LH induced dramatic transient increases in PG production by follicle wall and granulosa, but not theca, cells isolated from preovulatory follicles (0 h after GnRH). PG accumulation peaked on d 2 of culture, mimicking the secretion pattern after a gonadotropin surge in vivo. In cultures of follicle wall and granulosa cells, the PR antagonist mifepristone (MIFE, 1 microm) inhibited LH-induced PG secretion and the progestin medroxyprogesterone acetate (1 or 10 microm), but not the glucocorticoid dexamethasone (1 or 10 microm), overcame the effect of MIFE on PGs. Semiquantitative RT-PCR revealed that MIFE inhibited LH-induced expression of cyclooxygenase-2 mRNA in granulosa cells in vitro. Again, treatment with medroxyprogesterone acetate overcame the effect of MIFE. Together these results provide strong evidence that periovulatory increases in cyclooxygenase-2 mRNA, PGE, and PGF(2alpha) are mediated by gonadotropin-induced increases in progesterone/PR, indicating that in some species there is an important functional relationship between these pathways in the ovulatory cascade.     

An examination of cutting and other methods of DSH among children and adolescents presenting to an outpatient psychiatric clinic in New Zealand

This article explores the issue of cutting among children and adolescents presenting to an outpatient child and adolescent mental health service in New Zealand. Of interest is the extent to which children and adolescents who cut themselves also carry out other forms of deliberate self-harm (DSH). This study found that there was much cross over in the methods of DSH used and that cutters were not a distinguishable group. Therefore, those who present to mental health services with cutting need to be treated as carrying significant risk of fatality.     

Limiting and rationing treatment in paediatric and neonatal intensive care

In this chapter I consider the ethical decisions surrounding the provision and limitation of treatment offered to children requiring intensive care. I focus on the processes surrounding end of life decision making and consider how the concepts of futility, burden and uncertainty should impact upon these decisions. I also examine resource allocation to children's critical care services. The discussion does not provide a structure that will solve any given situation. It does take a practical approach to the issues faced by considering why we should engage in life limiting discussions; When they should occur; Who should be involved; How they should be carried out; and where and by what means withholding or withdrawal should occur. I have drawn the discussions closer to clinical practice with the intention of making them more useful, for those engaged in direct patient care, than those focused around philosophical principles.     

Cardiac hypertrophy, hypertrophic cardiomyopathy, and hyperparathyroidism--an association.

Left ventricular hypertrophy (symmetric, asymmetric, or hypertrophic cardiomyopathy) is an almost invariable accompaniment of primary hyperparathyroidism. Five of 18 patients with hypertrophic cardiomyopathy had raised serum concentrations of parathyroid hormone with normal serum calcium concentrations. Left ventricular hypertrophy did not occur in any of the six patients with hypercalcaemia alone. These relations suggest that parathyroid hormone rather than a rise in the extracellular calcium concentration is associated with a spectrum of left ventricular hypertrophy. All patients with increased circulating parathyroid hormone concentrations should have echocardiographic examination of the left ventricle. Conversely, parathyroid hormone concentrations should be measured in all patients with left ventricular hypertrophy from an unknown cause, especially those with hypertrophic cardiomyopathy.