Obligatory Role Research Articles

Side-Chain Proline-Based Polymers as Effective Inhibitors for In Vitro Aggregation of Insulin.

Insulin fibril formation is considered as the hallmark of several debilitating pathological conditions. To develop effective therapeutics that are able to control the amyloidogenesis process and inhibit fibril formation, herein we have designed a side-chain proline (Pro)-based homopolymer and block copolymers through the reversible addition-fragmentation chain transfer (RAFT) polymerization technique and further explored their obligatory role in the in vitro insulin fibrillation process. Using a variety of biophysical tools, including turbidity measurements, thioflavin T (ThT) fluorescence kinetics, tyrosine (Tyr) fluorescence study, Nile red (NR) fluorescence assay, dynamic light scattering (DLS) study, circular dichroism (CD) measurements, and isothermal titration calorimetry (ITC) techniques, we demonstrated that Pro-based polymers can significantly inhibit the insulin fibrillation process. Among them, the Pro-based homopolymer acts as the most potent inhibitor of insulin fibrillation as confirmed by ThT assay, CD study, and transmission electron microscopic (TEM) analysis. Tyrosine fluorescence measurements and NR fluorescence assay revealed that hydrophobic interactions are the crucial factor that mainly controls the inhibition process. Apart from hydrophobic interactions, polar interactions may also be responsible for the inhibition process as evaluated by ITC study.

Obligatory role for PKCδ in PIP2 -mediated activation of store-operated TRPC1 channels in vascular smooth muscle cells.

Key points In vascular smooth muscle cells (VSMCs), activation of Ca2+‐permeable store‐operated channels (SOCs) composed of canonical transient receptor potential channel 1 (TRPC1) subunits mediates Ca2+ entry pathways that regulate contraction, proliferation and migration, which are processes associated with vascular disease.Activation of TRPC1‐based SOCs requires protein kinase C (PKC) activity, which is proposed to phosphorylate TRPC1 proteins to promote channel opening by phosphatidylinositol 4,5‐bisphosphate (PIP2). We investigated the identity of the PKC isoform involved in activating TRPC1‐based SOCs in rat mesenteric artery VSMCs.TRPC1‐based SOCs were reduced by PKCδ inhibitors and knockdown of PKCδ expression. Store depletion induced interactions between TRPC1 and PKCδ and PKCδ‐dependent phosphorylation of TRPC1. Furthermore, generation of store‐operated interactions between PIP2 and TRPC1 and activation of TRPC1‐based SOCs by PIP2 required PKCδ.These findings reveal that PKCδ activity has an obligatory role in activating TRPC1‐based SOCs, through regulating PIP2‐mediated channel opening. In vascular smooth muscle cells (VMSCs), stimulation of Ca2+‐permeable canonical transient receptor potential channel 1 (TRPC1)‐based store‐operated channels (SOCs) mediates Ca2+ entry pathways that regulate cell contraction, proliferation and migration, which are processes associated with vascular disease. It is therefore important to understand how TRPC1‐based SOCs are activated. Stimulation of TRPC1‐based SOCs requires protein kinase C (PKC) activity, with store‐operated PKC‐dependent phosphorylation of TRPC1 essential for channel opening by phosphatidylinositol 4,5‐bisphosphate (PIP2). Experimental protocols used to activate TRPC1‐based SOCs suggest that the PKC isoform involved requires diacylglycerol (DAG) but is Ca2+‐insensitive, which are characteristics of the novel group of PKC isoforms (δ, ε, η, θ). Hence, the present study examined whether a novel PKC isoform(s) is involved in activating TRPC1‐based SOCs in contractile rat mesenteric artery VSMCs. Store‐operated whole‐cell cation currents were blocked by Pico145, a highly selective and potent TRPC1/4/5 channel blocker and T1E3, a TRPC1 blocking antibody. PKCδ was expressed in VSMCs, and selective PKCδ inhibitory peptides and knockdown of PKCδ expression with morpholinos oligomers inhibited TRPC1‐based SOCs. TRPC1 and PKCδ interactions and phosphorylation of TRPC1 induced by store depletion were both reduced by pharmacological inhibition and PKCδ knockdown. In addition, store‐operated PIP2 and TRPC1 interactions were blocked by PKCδ inhibition, and PKCδ was required for PIP2‐mediated activation of TRPC1 currents. These results identify the involvement of PKCδ in stimulation of TRPC1‐based SOCs and highlight that store‐operated PKCδ activity is obligatory for channel opening by PIP2, the probable activating ligand.

Ethical practice in the nursing profession: A normative analysis

ABSTRACT Background In response to an increasing number of litigations relating to nursing care errors, negligence or acts and omissions that arise mostly due to unprofessional or unethical behaviour compounded by the growing awareness of patient’s rights, nurse practitioners as such need an intervention by the regulatory body, the South African Nursing Council (SANC). Aim and purpose The argument presented pertains to the obligatory role of SANC as a regulatory body to uphold professional and ethical practice for nurses in terms of the nursing curriculum, the scope of practice, the code of ethics and continuing professional development. Methodology The methodology was based on the analysis of relevant SANC legislation under the provision of the Nursing Act 33 of 2005 as amended. Document review, a technique used to “categorise, investigate, interpret and identify limitations of written documents” was undertaken. The study referenced some of the general literature relating to nursing ethics and professionalism. A waiver for ethical clearance was obtained from the Human Research Ethics Committee. Conclusion The overall impact of the analysis lies in its contribution to enhancing nursing practice, with specific focus on upholding professional and ethical practice. SANC as a custodian of the nursing profession need to address limitations and develop a balance for the regulation and monitoring of ethical practice in both nursing education and clinical practice. Keywords: ethical practice; normative analysis; nursing profession; role; South African Nursing Council

Religion & Democracy: Interactions, Tensions, Possibilities

Religion & Democracy: Interactions, Tensions, Possibilities

Kv1.3 modulates neuroinflammation and neurodegeneration in Parkinson's disease.

Characterization of the key cellular targets contributing to sustained microglial activation in neurodegenerative diseases, including Parkinson's disease (PD), and optimal modulation of these targets can provide potential treatments to halt disease progression. Here, we demonstrated that microglial Kv1.3, a voltage-gated potassium channel, was transcriptionally upregulated in response to aggregated α-synuclein (αSynAgg) stimulation in primary microglial cultures and animal models of PD, as well as in postmortem human PD brains. Patch-clamp electrophysiological studies confirmed that the observed Kv1.3 upregulation translated to increased Kv1.3 channel activity. The kinase Fyn, a risk factor for PD, modulated transcriptional upregulation and posttranslational modification of microglial Kv1.3. Multiple state-of-the-art analyses, including Duolink proximity ligation assay imaging, revealed that Fyn directly bound to Kv1.3 and posttranslationally modified its channel activity. Furthermore, we demonstrated the functional relevance of Kv1.3 in augmenting the neuroinflammatory response by using Kv1.3-KO primary microglia and the Kv1.3-specific small-molecule inhibitor PAP-1, thus highlighting the importance of Kv1.3 in neuroinflammation. Administration of PAP-1 significantly inhibited neurodegeneration and neuroinflammation in multiple animal models of PD. Collectively, our results imply that Fyn-dependent regulation of Kv1.3 channels plays an obligatory role in accentuating the neuroinflammatory response in PD and identify Kv1.3 as a potential therapeutic target for PD.

TISSUE OXYGEN LEVEL IN ACUTE AND CHRONIC WOUND: A COMPARISON STUDY

Adequate oxygen supply to granulating tissue and its surroundings is considered as an elementary factor to render an optimum condition for proper wound healing. This paper focused on comparing the quantitative mean of transcutaneous oxygen saturation (StO2) between acute superficial wounds and chronic diabetic wounds in a randomized clinical study. A multispectral imaging system was employed for in-vivo measurement of wound StO2 across a visible wavelength range of nm. Processed StO2 maps of wounds using a reconstructed mathematical model revealed a mean StO2 of and , respectively, in acute and chronic wounds. The statistical test on the difference in the values between these two groups of wounds was shown to be statistically significant (). These crucial findings contribute to the existing knowledge on the StO2 range and its variation in different wound conditions. In conclusion, the proposed multispectral imaging system is able to produce clinical information of wound oxygenation level for a better understanding of the obligatory role of oxygen as a key determinant in healing outcomes.

Consider role of contractual obligations in decision‐making during times of crisis

The global crisis brought on by the COVID‐19 pandemic has forever changed higher education. In the beginning of the crisis, leaders on college campuses had to decide what to do with courses, students in residence halls, auxiliary services like food operations and bookstores, scheduled athletics competitions, and full‐time faculty and staff who normally reported to physical buildings. And those are just the obvious examples.

Dead Ringers? Legal Persons and the Deceased in European Data Protection Law

Notwithstanding suggestions that the treatment of legal and deceased person data during European data protection’s development has been broadly comparable, this paper finds that stark divergences are in fact apparent. Despite early fusion, legal persons have been increasingly seen to have lesser and, more importantly, qualitatively different information entitlements compared to natural persons, thereby leaving European data protection with a very limited and indirect role here. In contrast, natural persons and the deceased have not been conceived as normatively dichotomous and since the 1990s there has been growing interest both in establishing sui generis direct protection for deceased data and also indirect inclusion through a link with living natural persons. Whilst the case for some indirect inclusion is overwhelming, a broad approach to the inter-relational nature of data risks further destabilizing the personal data concept. Nevertheless, given that jurisdictions representing almost half of the EEA’s population now provide some direct protection and the challenges of managing digital data on death continue to grow, the time may be ripe for a ‘soft’ recommendation on direct protection in this area. Drawing on existing law and scholarship, such a recommendation could seek to specify the role of both specific control rights and diffuse confidentiality obligations, the criteria for time-limits in each case and the need for a balance with other rights and interests which recognises the significantly decreasing interest in protection over time.

Molecular insights into the inhibitory mechanisms of gallate moiety on the Aβ1–40 amyloid aggregation: A molecular dynamics simulation study

Alzheimer's disease is the most common form of neurodegenerative disease and the formation of Aβ amyloid aggregates has been widely demonstrated to be the principal cause of Alzheimer's disease. Our previous study and other studies suggested that the gallate moiety played an obligatory role in the inhibition process of naturally occurring polyphenols on Aβ amyloid fibrils formation. However, the detailed mechanisms were still unknown. Thus, in the present study, the gallic acid (GA) was specially selected and the molecular recognition mechanisms between GA molecules and Aβ1–40 monomer were examined and analyzed by molecular dynamics simulation. The in silico experiments revealed that GA significantly prevented the conformational changes of Aβ1–40 monomer with no β-sheet structure during the whole 100 ns. By analyzing the binding sites of GA molecules to Aβ1–40 monomer, we found that both hydrophilic and hydrophobic amino acid residues were participated in the binding of GA molecules to Aβ1–40 monomer. Moreover, results from the binding free energy analysis further demonstrated that the strength of polar interactions was significantly stronger than that of nonpolar interactions. We believed that our results could help to elucidate the underlying mechanisms of gallate moiety on the anti-amyloidogenic effects of polyphenols at the atomic level.

Contribution of KIR Channels and the Na+/K+‐ATPase to Local Vasodilation During Mild and Heavy Intensity Exercise in Humans

Potassium released from repolarizing skeletal muscle elicits vasodilation by stimulating inwardly‐rectifying K+ (KIR) channels and the Na+/K+‐ATPase on the vasculature, which can serve as a feedforward mechanism to increase muscle blood flow in proportion to the degree of muscle fiber recruitment during exercise. We tested the hypothesis that KIR channels and the Na+/K+‐ATPase play a greater role in the vasodilatory response to exercise at higher contraction intensities which require greater muscle fiber recruitment. Healthy volunteers performed mild and heavy intensity rhythmic handgrip exercise using a dynamic pulley system to lift a load equivalent to 10% or 25% of maximal voluntary contraction (MVC) at a rate of 20 contractions per minute. We measured muscle activation (electromyography; EMG), forearm blood flow (FBF; Doppler ultrasound) and mean arterial pressure (MAP; brachial artery catheter) to calculate vascular conductance (FVC; FBF/MAP*100) during 5 min of forearm exercise at each intensity. Responses were assessed during 1) control conditions, 2) local inhibition of either KIR channels alone (Protocol 1; intra‐arterial BaCl2) or the Na+/K+‐ATPase alone (Protocol 2; intra‐arterial ouabain; OUA), and 3) combined inhibition of KIR channels and the Na+/K+‐ATPase (Protocols 1 & 2; BaCl2 + OUA). As expected, muscle activation was greater during heavy compared to mild exercise (Control EMG, 65 ± 6 vs. 32 ± 3% of MVC; P < 0.05). In Protocol 1 (4 women, 5 men; 26 ± 1 yrs), BaCl2 reduced steady‐state FVC by −62 ± 21 ml/min/100 mmHg during mild exercise (Control: 170 ± 27 vs. BaCl2: 107 ± 10 ml/min/100 mmHg; P < 0.05), and combined inhibition with BaCl2 + OUA had no further effect (Δ FVC from Control, −67 ± 20; FVC, 103 ± 11 ml/min/100 mmHg; P = NS vs. BaCl2 alone). During heavy intensity exercise, BaCl2 reduced FVC by −59 ± 13 ml/min/100 mmHg (Control: 333 ± 24 vs. BaCl2: 274 ± 29 ml/min/100 mmHg; P < 0.05), which was similar to the effect of BaCl2 during mild exercise (Δ FVC P = NS vs. mild exercise). However, in contrast to mild exercise, combined inhibition with BaCl2 + OUA further attenuated FVC during heavy intensity exercise (Δ FVC from Control, □108 ± 24; FVC, 225 ± 15 ml/min/100 mmHg; P < 0.05 vs. BaCl2). In Protocol 2 (2 women, 3 men; 26 ± 2 yrs), OUA alone had no effect on FVC during mild exercise (Δ FVC from Control, −2 ± 6; FVC, Control: 141 ± 31 vs. OUA: 139 ± 36; P = NS) or heavy exercise (Δ FVC from Control, −11 ± 11; FVC, Control: 325 ± 70 vs. OUA: 313 ± 71; P = NS), whereas combined inhibition with BaCl2 + OUA reduced FVC to a similar extent as in Protocol 1 (mild exercise, Δ FVC from Control: −43 ± 4; FVC: 98 ± 30; heavy exercise, Δ FVC from Control: −109 ± 16; FVC: 216 ± 66 ml/min/100 mmHg, both P < 0.05 vs. Control and OUA alone). We conclude that KIR channel activation contributes similarly to skeletal muscle vascular tone at mild and heavy exercise intensities. Furthermore, an obligatory role for Na+/K+‐ATPase in vascular control is not revealed until KIR channels are inhibited during heavy intensity exercise.Support or Funding InformationNIH R01‐HL119337

Gnotobiotic rats reveal an obligatory role of microbiota in blood pressure

Blood pressure is a cardinal vital sign that informs clinicians for both acute and long‐term clinical care. Elevated blood pressure or hypertension is the single largest risk factor for cardiovascular diseases which are the leading cause of human deaths. Current clinical management of blood pressure is focused on restoring homeostasis of the host alone, which targets an incomplete holobiont i.e., without accounting for commensal gut microbiota. Recent evidence from the CARDIA study in humans and multiple studies using animal models suggest that development of hypertension in the host is associated with alterations in microbiotal communities. Whether microbiota is a required component of the holobiont to maintain cardiovascular health is unknown. Here we examined whether microbiota is necessary for blood pressure and vascular homeostasis by functional evaluation of the gut homeostasis, hemodynamic, and vascular function of gnotobiotic rats reconstituted with microbiota to represent the complete holobiont. Gnotobiotic rats were used to represent incomplete holobionts. To reconstitute complete holobionts, gnotobiotic rats were co‐housed with conventionally‐raised rats. Acquisition of microbiota was evaluated through monitoring of gross ceca and fecal samples by metagenomic 16S sequencing. BP was recorded and vascular, renal, hepatic, cardiac and gut features were assessed using histology and ex vivo myography. Markers of innate immune effectors (Immune cell population, level of Lcn2, Gut permeability) were used to examine the nature and extent of host immune cell processes concomitantly occurring along with observations of host hemodynamics. Compared to the reconstituted holobiont represented by the animals exposed to microbiota, the incomplete‐holobiont represented by gnotobiotic rats, had significantly lower BP (SBP of germ free:109±8 mmHg, SBP of conventionalized:138±10mmHg*) and vascular contractility responses to phenylephrine (Emax (mN): germ‐free: 6.9±1.3, GFC: 11.7±0.7*). Acute exposure of the host to microbiota reconstituted gut microbiotal communities, significantly boosted their gut epithelial cell proliferation, innate immune function and restored vascular contractility. These data indicate that in addition to the dependency of the host on microbiota for essential bodily functions such as digestion of plant‐derived complex carbohydrates, the host is also dependent on microbiota for maintaining blood pressure and vascular functionSupport or Funding InformationNational Institutes of Health (R01HL143082)

Voluntary assisted dying: should conscientious objection be unconditional?

This paper argues that although legislation on voluntary assisted dying should always allow medical practitioners to practise conscientious objection, there should always be an obligation on the conscientious objector to refer anyone who inquires about voluntary assisted dying to other medical practitioners who have no conscientious objection. The paper refutes the concern that this requirement effectively undermines the conscientious objector's right to object, explaining exactly why this concern is misplaced.

Ventilatory acclimatization to hypoxia: Time to express a critical central message.

Ventilatory acclimatization to hypoxia: Time to express a critical central message.

Photorhabdus: a tale of contrasting interactions.

Different model systems have, over the years, contributed to our current understanding of the molecular mechanisms underpinning the various types of interaction between bacteria and their animal hosts. The genus Photorhabdus comprises Gram-negative insect pathogenic bacteria that are normally found as symbionts that colonize the gut of the infective juvenile stage of soil-dwelling nematodes from the family Heterorhabditis. The nematodes infect susceptible insects and release the bacteria into the insect haemolymph where the bacteria grow, resulting in the death of the insect. At this stage the nematodes feed on the bacterial biomass and, following several rounds of reproduction, the nematodes develop into infective juveniles that leave the insect cadaver in search of new hosts. Therefore Photorhabdus has three distinct and obligate roles to play during this life-cycle: (1) Photorhabdus must kill the insect host; (2) Photorhabdus must be capable of supporting nematode growth and development; and (3) Photorhabdus must be able to colonize the gut of the next generation of infective juveniles before they leave the insect cadaver. In this review I will discuss how genetic analysis has identified key genes involved in mediating, and regulating, the interaction between Photorhabdus and each of its invertebrate hosts. These studies have resulted in the characterization of several new families of toxins and a novel inter-kingdom signalling molecule and have also uncovered an important role for phase variation in the regulation of these different roles.

Protecting children’s rights under the ECHR: the role of positive obligations

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How Does Place Matter to Highly Skilled Migrants?: <i>Work/non-work experiences of international physicians in Norway and Sweden</i>

The article tackles the question of how place matters to migrant physicians in the regions of Agder in Norway and Skane in Sweden by exploring how place-specific conditions affect their experiences in the work, private, family and social domains of life. For this purpose, the article uses thematic analysis of the narrative material gathered through 25 semi-structured interviews. The lens of work/non-work domains, combined with a practice-oriented approach to place, highlights the complexity of lived experiences as they evolve in a particular context. Three main findings are identified: the non-homogenous significance of place across life domains, the vital role of transborder connections and obligations that affect individual and family resources for work/non-work negotiations in the place of settlement and the limits to the skill-based privileges in the place of settlement, which are notable in the domain of work but not replicated in non-work domains.

Dynamic Recovery from Depression Enables Rate Encoding in Inhibitory Synapses.

SummaryParvalbumin-expressing fast-spiking interneurons (PV-INs) control network firing and the gain of cortical response to sensory stimulation. Crucial for these functions, PV-INs can sustain high-frequency firing with no accommodation. However, PV-INs also exhibit short-term depression (STD) during sustained activation, largely due to the depletion of synaptic resources (vesicles). In most synapses the rate of replenishment of depleted vesicles is constant, determining an inverse relationship between depression levels and the activation rate, which theoretically, severely limits rate-coding capabilities. We examined STD of the PV-IN to pyramidal cell synapse in the mouse visual cortex and found that in these synapses the recovery from depression is not constant but increases linearly with the frequency of use. By combining modeling, dynamic clamp, and optogenetics, we demonstrated that this recovery enables PV-INs to reduce pyramidal cell firing in a linear manner, which theoretically is crucial for controlling the gain of cortical visual responses.

Acoustic tracheal rupture provides insights into larval mosquito respiration

Acoustic larviciding (AL) occurs by exposing mosquito larvae to acoustic energy that ruptures their dorsal tracheal trunks (DTTs) by the expulsion of gas bubbles into the body. In studying this technique, we serendipitously identified undescribed anatomical and physiological respiratory features. The classical theory of respiration is that the siphon and DTTs play obligate roles in respiration. Our results contradict the accepted theory that culicine larvae respire via atmospheric gas exchange. We identified an undescribed tracheal occlusion (TO) at the posterior extremities the DTTs. The TOs appear necessary for the acoustic rupture of DTTs; this constriction prevents the escape of energized gas from the siphon and allows the tracheal system to be pressurized. With a pressurized isolated tracheal system, metabolic gas exchange directly with the atmosphere is unlikely and could mostly occur through the chitin and setae. Future studies are needed to explore respiration and elucidate the mechanisms of oxygen absorption and carbon dioxide elimination.

Adherens Junctions: Guardians of Cortical Development.

Apical radial glia comprise the pseudostratified neuroepithelium lining the embryonic lateral ventricles and give rise to the extensive repertoire of pyramidal neuronal subtypes of the neocortex. The establishment of a highly apicobasally polarized radial glial morphology is a mandatory prerequisite for cortical development as it governs neurogenesis, neural migration and the integrity of the ventricular wall. As in all epithelia, cadherin-based adherens junctions (AJs) play an obligate role in the maintenance of radial glial apicobasal polarity and neuroepithelial cohesion. In addition, the assembly of resilient AJs is critical to the integrity of the neuroepithelium which must resist the tensile forces arising from increasing CSF volume and other mechanical stresses associated with the expansion of the ventricles in the embryo and neonate. Junctional instability leads to the collapse of radial glial morphology, disruption of the ventricular surface and cortical lamination defects due to failed neuronal migration. The fidelity of cortical development is therefore dependent on AJ assembly and stability. Mutations in genes known to control radial glial junction formation are causative for a subset of inherited cortical malformations (neuronal heterotopias) as well as perinatal hydrocephalus, reinforcing the concept that radial glial junctions are pivotal determinants of successful corticogenesis. In this review we explore the key animal studies that have revealed important insights into the role of AJs in maintaining apical radial glial morphology and function, and as such, have provided a deeper understanding of the aberrant molecular and cellular processes contributing to debilitating cortical malformations. We highlight the reciprocal interactions between AJs and the epithelial polarity complexes that impose radial glial apicobasal polarity. We also discuss the critical molecular networks promoting AJ assembly in apical radial glia and emphasize the role of the actin cytoskeleton in the stabilization of cadherin adhesion – a crucial factor in buffering the mechanical forces exerted as a consequence of cortical expansion.

Shared Obligations and the Responsibility of an International Organization and its Member States: The Case of EU Mixed Agreements

Discussions on the allocation of international responsibility between an international organization and its member states do not comprehensively engage with the role of obligations in assigning responsibility to the organization and/or its members. The present article sets out what will be termed an obligations-based approach to the allocation of international responsibility by exploring the phenomenon of sharing international obligations by an international organization and its members, as well as the implications thereof for their responsibility under international law. It will do so by focusing on the practice of concluding mixed agreements by the EU and its member states, which commonly results in overlapping obligations for the organization and its members. It is ultimately argued that a distinction should be made between two types of shared obligations in mixed agreements in order to untangle who can be held responsible in case of a breach: the EU, the member state(s), or both.