Dynamic Evolution of Bacterial Ligand Recognition by Formyl Peptide Receptors.

The detection of invasive pathogens is critical for host immune defense. Cell surface receptors play a key role in the recognition of diverse microbe-associated molecules, triggering leukocyte recruitment, phagocytosis, release of antimicrobial compounds, and cytokine production. The intense evolutionary forces acting on innate immune receptor genes have contributed to their rapid diversification across plants and animals. However, the functional consequences of immune receptor divergence are often unclear. Formyl peptide receptors (FPRs) comprise a family of animal G protein-coupled receptors which are activated in response to a variety of ligands including formylated bacterial peptides, pathogen virulence factors, and host-derived antimicrobial peptides. FPR activation in turn promotes inflammatory signaling and leukocyte migration to sites of infection. Here we investigate patterns of gene loss, diversification, and ligand recognition among FPRs in primates and carnivores. We find that FPR1, which plays a critical role in innate immune defense in humans, has been lost in New World primates. Amino acid variation in FPR1 and FPR2 among primates and carnivores is consistent with a history of repeated positive selection acting on extracellular domains involved in ligand recognition. To assess the consequences of FPR divergence on bacterial ligand interactions, we measured binding between primate FPRs and the FPR agonist Staphylococcus aureus enterotoxin B, as well as S. aureus FLIPr-like, an FPR inhibitor. We found that few rapidly evolving sites in primate FPRs are sufficient to modulate recognition of bacterial proteins, demonstrating how natural selection may serve to tune FPR activation in response to diverse microbial ligands.

Barriers and facilitators to implementing pharmacist-provided comprehensive medication management in primary care transformation.

PURPOSE: How to effectively integrate pharmacists into team-based models of care to maximize the benefit they bring to patients and care teams, especially during times of primary care transformation (PCT), remains unknown. The objective of this study was to identify barriers and facilitators when integrating pharmacist-provided comprehensive medication management (CMM) services into a health system's team-based PCT using the Consolidated Framework for Implementation Research (CFIR). METHODS: Semistructured qualitative interviews were carried out with 22 care team members regarding their perceptions of the implementation of CMM in the PCT. Transcripts were coded to identify CMM implementation barriers and facilitators, and resulting codes were mapped to corresponding CFIR domains and constructs. RESULTS: Fifteen codes emerged that were labeled as either a barrier or a facilitator to implementing CMM in the PCT. Facilitators were the perception of CMM as an invaluable resource, precharting, tailored appointment lengths, insurance coverage, increased pharmacy presence, enhanced team-based care, location of CMM, and identification of CMM advocates. Barriers included limited clinic leadership involvement, a need for additional resources, CMM pharmacists not always feeling part of the core team, understanding of and training around CMM's role in the PCT, changing mindsets to utilize resources such as CMM more frequently, underutilization of CMM, and CMM scheduling. CONCLUSION: Clinical pharmacists providing CMM represent a valuable interdisciplinary care team member who can help improve healthcare quality and access to primary care. Identifying and addressing implementation barriers and facilitators early during PCT rollout is critical to the success of team-based services such as CMM and becoming a learning health system.

Delivering Care to High-Cost High-Need Patients: Lessons Learned in the Development of a Complex Care Primary Care Team.

As part of a population health-focused primary care transformation, in 2019 a health system in Minnesota developed a primary care team to exclusively care for high-cost high-need patients. Through its development and implementation, the team has discovered several key lessons in delivering care to complex patients. These lessons include the benefits of more integrative team-based care, the need and advantages of designated complex care team members, the importance of teamwork both within and outside of the complex care team, the need for frequent communication, and the importance of identifying mental health needs. In addition, there are several areas that require ongoing research and exploration, such as determining when a patient is able to graduate out of the program, how to enhance access to the complex care team, determining appropriate visit characteristics, and model feasibility. While addressing the needs of high cost high need patients is essential to improving quality of care and decreasing health care costs, there are several unique challenges and opportunities that come with caring for this patient population. Although this highly integrated model of care continues to evolve, the initial lessons learned may inform other health systems and care teams undertaking the care of complex patients.

Diversification of CD1 Molecules Shapes Lipid Antigen Selectivity.

Molecular studies of host-pathogen evolution have largely focused on the consequences of variation at protein-protein interaction surfaces. The potential for other microbe-associated macromolecules to promote arms race dynamics with host factors remains unclear. The cluster of differentiation 1 (CD1) family of vertebrate cell surface receptors plays a crucial role in adaptive immunity through binding and presentation of lipid antigens to T-cells. Although CD1 proteins present a variety of endogenous and microbial lipids to various T-cell types, they are less diverse within vertebrate populations than the related major histocompatibility complex (MHC) molecules. We discovered that CD1 genes exhibit a high level of divergence between simian primate species, altering predicted lipid-binding properties and T-cell receptor interactions. These findings suggest that lipid-protein conflicts have shaped CD1 genetic variation during primate evolution. Consistent with this hypothesis, multiple primate CD1 family proteins exhibit signatures of repeated positive selection at surfaces impacting antigen presentation, binding pocket morphology, and T-cell receptor accessibility. Using a molecular modeling approach, we observe that interspecies variation as well as single mutations at rapidly-evolving sites in CD1a drastically alter predicted lipid binding and structural features of the T-cell recognition surface. We further show that alterations in both endogenous and microbial lipid-binding affinities influence the ability of CD1a to undergo antigen swapping required for T-cell activation. Together these findings establish lipid-protein interactions as a critical force of host-pathogen conflict and inform potential strategies for lipid-based vaccine development.

Development, Testing and Results of a Patient Medication Experience Documentation Tool for Use in Comprehensive Medication Management Services.

The medication experience is an individual's subjective experience of taking a medication in daily life and can be at the root of drug therapy problems. It is recommended that the patient-centered approach to comprehensive medication management (CMM) starts with an understanding of the patient's medication experience. This study aims to develop a medication experience documentation tool for use in CMM services, and to understand the usefulness and challenges of using the tool in practice. The tool was developed based on previous research on patients' medication experiences. It was tested in two rounds by ten CMM pharmacists utilizing the tool as they provided care to patients. Focus groups were conducted to revise the tool after each round and to understand pharmacists' experiences. The tool was tested for 15 weeks in 407 patient encounters. There was at least one medication experience documented in the electronic medical record 62% of the time. Pharmacists found the tool helpful in raising awareness of the medication experience and motivational interviewing strategies, planning for follow-up visits, as a teaching tool, and making pharmacists realize the fluidity of the medication experience. The tool offered pharmacists a better way to recognize and address medication experiences affecting medication taking behaviors.

Activation of Discs large by aPKC aligns the mitotic spindle to the polarity axis during asymmetric cell division.

Asymmetric division generates cellular diversity by producing daughter cells with different fates. In animals, the mitotic spindle aligns with Par complex polarized fate determinants, ensuring that fate determinant cortical domains are bisected by the cleavage furrow. Here, we investigate the mechanisms that couple spindle orientation to polarity during asymmetric cell division of Drosophila neuroblasts. We find that the tumor suppressor Discs large (Dlg) links the Par complex component atypical Protein Kinase C (aPKC) to the essential spindle orientation factor GukHolder (GukH). Dlg is autoinhibited by an intramolecular interaction between its SH3 and GK domains, preventing Dlg interaction with GukH at cortical sites lacking aPKC. When co-localized with aPKC, Dlg is phosphorylated in its SH3 domain which disrupts autoinhibition and allows GukH recruitment by the GK domain. Our work establishes a molecular connection between the polarity and spindle orientation machineries during asymmetric cell division.

In preeclampsia, maternal third trimester subcutaneous adipocyte lipolysis is more resistant to suppression by insulin than in healthy pregnancy.

Obesity increases preeclampsia risk, and maternal dyslipidemia may result from exaggerated adipocyte lipolysis. We compared adipocyte function in preeclampsia with healthy pregnancy to establish whether there is increased lipolysis. Subcutaneous and visceral adipose tissue biopsies were collected at caesarean section from healthy (n=31) and preeclampsia (n=13) mothers. Lipolysis in response to isoproterenol (200 nmol/L) and insulin (10 nmol/L) was assessed. In healthy pregnancy, subcutaneous adipocytes had higher diameter than visceral adipocytes (P<0.001). Subcutaneous and visceral adipocyte mean diameter in preeclampsia was similar to that in healthy pregnant controls, but cell distribution was shifted toward smaller cell diameter in preeclampsia. Total lipolysis rates under all conditions were lower in healthy visceral than subcutaneous adipocytes but did not differ after normalization for cell diameter. Visceral adipocyte insulin sensitivity was lower than subcutaneous in healthy pregnancy and inversely correlated with plasma triglyceride (r=-0.50; P=0.004). Visceral adipose tissue had lower ADRB3, LPL, and leptin and higher insulin receptor messenger RNA expression than subcutaneous adipose tissue. There was no difference in subcutaneous adipocyte lipolysis rates between preeclampsia and healthy controls, but subcutaneous adipocytes had lower sensitivity to insulin in preeclampsia, independent of cell diameter (P<0.05). In preeclampsia, visceral adipose tissue had higher LPL messenger RNA expression than subcutaneous. In conclusion, in healthy pregnancy, the larger total mass of subcutaneous adipose tissue may release more fatty acids into the circulation than visceral adipose tissue. Reduced insulin suppression of subcutaneous adipocyte lipolysis may increase the burden of plasma fatty acids that the mother has to process in preeclampsia.

Improved pregnancy rates with luteinizing hormone supplementation in patients undergoing ovarian stimulation for IVF.

INTRODUCTION: Luteinizing hormone (LH) is believed to play a role in follicle maturation during the natural cycle. However, the need for co-treatment with recombinant LH (rLH) for controlled ovarian stimulation is controversial. PURPOSE: The primary objective of our study was to determine if pregnancy rates are improved when rLH is used in addition to rFSH for ovarian stimulation. Secondary outcomes were fertilization rate, implantation rate and live birth rate. METHODS: A retrospective cohort study was performed of 1565 IVF or ICSI cycles. Outcomes were compared between ovarian stimulation cycles from 2007 when rLH and rFSH was used (n = 765) to 2006 when rFSH only was used (n-800). RESULTS: Improved outcomes were found for rLH + rFSH versus rFSH alone for; pregnancy rate (61% and 54% respectively, p = 0.006), live birth rate (49% and 42% respectively, P = 0.01), fertilization rate (74% versus 72% respectively, p = 0.04 and implantation rate (41% versus 37% respectively, p = 0.03). CONCLUSIONS: Our large retrospective cohort study showed an improved pregnancy rate and live birth rate with rLH supplementation. This was associated with an improved fertilization and implantation rate and therefore may reflect an improvement in oocyte quality and/or uterine receptivity.

The effects of modafinil on mood and cognition in Huntington's disease.

RATIONALE: The wake-promoting agent modafinil selectively improves neuropsychological task performance in healthy volunteers, in adults with attention deficit hyperactivity disorder (ADHD) and in schizophrenia. We examined whether modafinil induced similar effects in individuals with Huntington's disease (HD). MATERIALS AND METHODS: Twenty patients with genetically proven, mild HD participated in a double-blind, randomised, placebo-controlled cross-over study using a single 200 mg dose of modafinil. Patients undertook a battery of neuropsychological tests including measures of cognition and mood. RESULTS: Modafinil increased alertness as indexed by visual analogue scales. Modafinil did not elicit any significant improvements in cognitive function or mood. Modafinil had a deleterious effect on visual recognition and working memory. CONCLUSIONS: Two hundred milligrams acute modafinil administration did improve alertness but did not improve cognition or mood in patients with mild HD. A multiple dose, chronic administration study is needed before the potential clinical utility of modafinil in HD is discounted.

Cerebral blood flow and ventilatory sensitivity to CO2 measured with the modified rebreathing method.

The ventilatory response to carbon dioxide (CO2) measured by modified rebreathing (SrVE) is closer to that measured by the steady-state method (SsVE) than is the response measured by Read's rebreathing method. Furthermore, the value estimated by the steady-state method depends upon the number of data points used to measure it. We planned to assess if these observations were also true for cerebral blood flow (CBF), as measured by steady-state (SsCBF) and modified rebreathing (SrCBF) tests. Six subjects undertook two protocols, one in the steady-state and one with modified rebreathing. SsVE depended upon the number of data points used to calculate it, and SsVE and SrVE were similar. However, this was not the case with SsCBF, and SsCBF was much higher than SrCBF. These findings are consistent with the notions that the specific CO2 stimulus differs for CBF control as compared with ventilation (VE) control, and that prior hypocapnia has an effect on CBF and VE for longer than the duration of the hypocapnia.

Cerebral blood flow sensitivities to CO2 measured with steady-state and modified rebreathing methods.

It is well established that the ventilatory response to carbon dioxide (CO(2)) measured by modified rebreathing (Sr(VE)) is closer to that measured by the steady-state method (Ss(VE)) than is the response measured by Read's rebreathing method. It is also known that the value estimated by the steady-state method depends upon the combination of data points used to measure it. The aim of this study was to investigate if these observations were also true for cerebral blood flow (CBF), as measured by steady-state (Ss(CBF)) and modified rebreathing (Sr(CBF)) tests. Six subjects undertook two protocols: (a) steady state: PET(CO2) was held at 1.5 mm Hg above normal (isocapnia) for 10 min, then raised to three levels of hypercapnia, (8 min each; 6.5, 11.5 and 16.5 mm Hg above normal, separated by 4 min isocapnia). End-tidal PO2 was held at 300 mm Hg; (b) modified rebreathing: subjects underwent 6 min of voluntary hyperventilation to PET(CO2) approximately 20 mm Hg, and then rebreathed via a 6l bag filled with 6.5% CO(2) in O(2). We confirmed that the value for Ss(VE) depended upon the combination of data points used to calculate it, and also confirmed that Ss(VE) and Sr(VE) were similar. However, this was not the case with CBF. Estimates of Ss(CBF) were the same, regardless of the data points used in calculation, and Ss(CBF) was 89% greater than Sr(CBF) (P<0.05). We interpret these findings as consistent with the notion that the specific CO(2) stimulus differs for CBF and ventilatory control. The data also indicate that prior hypocapnia in the modified rebreathing protocol may have a persistent effect on both cerebral vessels and central ventilatory control.

Effects of prior heavy-intensity exercise during single-leg knee extension on VO2 kinetics and limb blood flow.

The effects of prior heavy-intensity exercise on O(2) uptake (Vo(2)) kinetics of a second heavy exercise may be due to vasodilation (associated with metabolic acidosis) and improved muscle blood flow. This study examined the effect of prior heavy-intensity exercise on femoral artery blood flow (Qleg) and its relationship with Vo(2) kinetics. Five young subjects completed five to eight repeats of two 6-min bouts of heavy-intensity one-legged, knee-extension exercise separated by 6 min of loadless exercise. Vo(2) was measured breath by breath. Pulsed-wave Doppler ultrasound was used to measure Qleg. Vo(2) and blood flow velocity data were fit using a monoexponential model to identify phase II and phase III time periods and estimate the response amplitudes and time constants (tau). Phase II Vo(2) kinetics was speeded on the second heavy-intensity exercise [mean tau (SD), 29 (10) s to 24 (10) s, P < 0.05] with no change in the phase II (or phase III) amplitude. Qleg was elevated before the second exercise [1.55 (0.34) l/min to 1.90 (0.25) l/min, P < 0.05], but the amplitude and time course [tau, 25 (13) s to 35 (13) s] were not changed, such that throughout the transient the Qleg (and DeltaQleg/DeltaVo(2)) did not differ from the prior heavy exercise. Thus Vo(2) kinetics were accelerated on the second exercise, but the faster kinetics were not associated with changes in Qleg. Thus limb blood flow appears not to limit Vo(2) kinetics during single-leg heavy-intensity exercise nor to be the mechanism of the altered Vo(2) response after heavy-intensity prior exercise.

Kinetics of .VO2 and femoral artery blood flow during heavy-intensity, knee-extension exercise.

It has been suggested that, during heavy-intensity exercise, O(2) delivery may limit oxygen uptake (.VO2) kinetics; however, there are limited data regarding the relationship of blood flow and .VO2 kinetics for heavy-intensity exercise. The purpose was to determine the exercise on-transient time course of femoral artery blood flow (Q(leg)) in relation to .VO2 during heavy-intensity, single-leg, knee-extension exercise. Five young subjects performed five to eight repeats of heavy-intensity exercise with measures of breath-by-breath pulmonary .VO2 and Doppler ultrasound femoral artery mean blood velocity and vessel diameter. The phase 2 time frame for .VO2 and Q(leg) was isolated and fit with a monoexponent to characterize the amplitude and time course of the responses. Amplitude of the phase 3 response was also determined. The phase 2 time constant for .VO2 of 29.0 s and time constant for Q(leg) of 24.5 s were not different. The change (Delta) in .VO2 response to the end of phase 2 of 0.317 l/min was accompanied by a DeltaQ(leg) of 2.35 l/min, giving a DeltaQ(leg)-to-Delta.VO2 ratio of 7.4. A slow-component .VO2 of 0.098 l/min was accompanied by a further Q(leg) increase of 0.72 l/min (DeltaQ(leg)-to-Delta.VO2 ratio = 7.3). Thus the time course of Q(leg) was similar to that of muscle .VO2 (as measured by the phase 2 .VO2 kinetics), and throughout the on-transient the amplitude of the Q(leg) increase achieved (or exceeded) the Q(leg)-to-.VO2 ratio steady-state relationship (ratio approximately 4.9). Additionally, the .VO2 slow component was accompanied by a relatively large rise in Q(leg), with the increased O(2) delivery meeting the increased Vo(2). Thus, in heavy-intensity, single-leg, knee-extension exercise, the amplitude and kinetics of blood flow to the exercising limb appear to be closely linked to the .VO2 kinetics.

Peripheral chemoreceptor control of ventilation following sustained hypoxia in young and older adult humans.

The rate and duration of peripheral chemoreceptor resensitization following sustained hypoxia was characterized in young and older (74-year-old) adults. In addition, cerebral blood velocity (CBV) was measured in young subjects during and following the relief from sustained hypoxia. Following 20 min of sustained eucapnic hypoxia (50 mmHg), subjects were re-exposed to brief (1.5 min) hypoxic pulses (50 mmHg), and the magnitude of the ventilatory response was used to gauge peripheral chemosensitivity. Five minutes after the relief from sustained hypoxia, ventilation (V(E)) increased to 40.3 +/- 4.5% of the initial hypoxic ventilatory response, and by 36 min V(E) increased to 100%, indicating that peripheral chemosensitivity to hypoxia was restored. The V(E) response magnitude plotted versus time demonstrated that V(E), hence peripheral chemosensitivity, was restored at a rate of 1.9% per minute. Cerebral blood flow (CBF, inferred from CBV) remained constant during sustained hypoxia and increased by the same magnitude during the hypoxic pulses, suggesting that CBF has a small, if any, impact on the decline in V(E) during hypoxia and its subsequent recovery. To address the issue of whether hypoxic pulses affect subsequent challenges, series (continuous hypoxic pulses at various recovery intervals) and parallel (only 1 pulse per trial) methods were used. There were no differences in the ventilatory responses between the series and parallel methods. Older adults demonstrated a similar rate of recovery as in the young, suggesting that ageing in active older adults does not affect the peripheral chemoreceptor response.

Cerebral blood flow sensitivity to CO2 measured with steady-state and Read's rebreathing methods.

The ventilatory response to carbon dioxide (CO2) measured by the steady-state method is lower than that measured by Read's rebreathing method. A change in end-tidal P CO2 (PET CO2) results in a lower increment change in brain tissue P CO2 (Pt CO2) in the steady-state than with rebreathing: since Pt(CO2) determines the ventilatory response to CO2, the response is lower in the steady-state. If cerebral blood flow (CBF) responds to Pt CO2, the CBF-CO2 response should be lower in the steady-state than with rebreathing. Six subjects undertook two protocols, (a) steady-state: PET CO2 was held at 1.5 mmHg above normal (isocapnia) for 10 min, then raised to three levels of hypercapnia, (8 min each; 6.5, 11.5 and 16.5 mmHg above normal, separated by 4 min isocapnia). End-tidal P O2 was held at 300 mmHg; (b) rebreathing: subjects rebreathed via a 6 L bag filled with 6.5% CO2 in O2. Transcranial Doppler-derived CBF yielded a higher CBF-CO2 sensitivity in the steady-state than with rebreathing, suggesting that CBF does not respond to Pt CO2.