The acyltransferase Gpc1 is both a target and an effector of the unfolded protein response in Saccharomyces cerevisiae.

The unfolded protein response (UPR) is sensitive to proteotoxic and membrane bilayer stress, both of which are sensed by the ER protein Ire1. When activated, Ire1 splices HAC1 mRNA, producing a transcription factor that targets genes involved in proteostasis and lipid metabolism, among others. The major membrane lipid phosphatidylcholine (PC) is subject to phospholipase-mediated deacylation, producing glycerophosphocholine (GPC), followed by reacylation of GPC through the PC deacylation/reacylation pathway (PC-DRP). The reacylation events occur via a two-step process catalyzed first by the GPC acyltransferase Gpc1, followed by acylation of the lyso-PC molecule by Ale1. However, whether Gpc1 is critical for ER bilayer homeostasis is unclear. Using an improved method for C14-choline-GPC radiolabeling, we first show that loss of Gpc1 results in abrogation of PC synthesis through PC-DRP and that Gpc1 colocalizes with the ER. We then probe the role of Gpc1 as both a target and an effector of the UPR. Exposure to the UPR-inducing compounds tunicamycin, DTT, and canavanine results in a Hac1-dependent increase in GPC1 message. Further, cells lacking Gpc1 exhibit increased sensitivity to those proteotoxic stressors. Inositol limitation, known to induce the UPR via bilayer stress, also induces GPC1 expression. Finally, we show that loss of GPC1 induces the UPR. A gpc1Δ mutant displays upregulation of the UPR in strains expressing a mutant form of Ire1 that is unresponsive to unfolded proteins, indicating that bilayer stress is responsible for the observed upregulation. Collectively, our data indicate an important role for Gpc1 in yeast ER bilayer homeostasis.

US Nationwide Multi-City Media Coverage of COVID-19 Responses: Community Structure Theory, Belief System, and a "Violated Way of Life".

Community structure analysis compared city characteristics and newspaper coverage of state/local government responses to COVID-19 in 25 major U.S. cities, sampling all 250+ word articles from 4/4/20 to 7/6/20. The resulting 588 articles were coded for "prominence" and "direction" (favorable/unfavorable/balanced-neutral coverage), then combined into each newspaper's composite "Media Vector" (range=0.3552 to -0.5197, or 0.8749). Twenty-one of 25 newspapers (84%) displayed unfavorable coverage of local COVID-19 responses. Pearson correlations and regression analysis confirmed a muscular "violated way of life" pattern, when a community perceives itself as threatened by a "biological threat or a threat to a cherished way of life." Political and belief system polarization (in particular percent Evangelical and percent voting Republican) were strongly associated with unfavorable coverage of local pandemic responses, compared to more favorable responses linked to percent voting Democratic or percent Catholic. Vulnerability (percent uninsured) was also linked to negative coverage. Conversely, two different measures of access to healthcare (percent municipal spending on health and welfare, and physicians/100,000) were significantly linked to favorable coverage of the same local government efforts. Community structure theory's grass roots "bottom up" expectations linking community demographics to variations in reporting on critical issues were robustly confirmed.

Assessing IRS performance in a gender-integrated vector control programme on Bioko Island, Equatorial Guinea, 2010-2021.

BACKGROUND: Indoor residual spraying (IRS) is a common vector control strategy in countries with high malaria burden. Historically, social norms have prevented women from working in IRS programmes. The Bioko Island Malaria Elimination Project has actively sought to reduce gender inequality in malaria control operations for many years by promoting women's participation in IRS. METHODS: This study investigated the progress of female engagement and compared spray productivity by gender from 2010 to 2021, using inferential tests and multivariable regression. Spray productivity was measured by rooms sprayed by spray operator per day (RSOD), houses sprayed by spray operator per day (HSOD), and the daily productivity ratio (DPR), defined as the ratio of RSOD to HSOD, which standardized productivity by house size. RESULTS: The percentage of women participating in IRS has increased over time. The difference in DPR comparing male and female spray operators was only statistically significant (p < 0.05) for two rounds, where the value was higher for women compared to men. Regression analyses showed marginal, significant differences in DPR between men and women, but beta coefficients were extremely small and thus not indicative of a measurable effect of gender on operational performance. CONCLUSIONS: The quantitative analyses of spray productivity are counter to stigmatizing beliefs that women are less capable than male counterparts during IRS spray rounds. The findings from this research support the participation of women in IRS campaigns, and a renewed effort to implement equitable policies and practices that intentionally engage women in vector control activities.

RNA-Seq Reveals Sex Differences in Gene Expression during Peripheral Neuropathic Inflammation and in Pain Relief from a COX-2 Inhibiting Theranostic Nanoemulsion.

Given decades of neuroinflammatory pain research focused only on males, there is an urgent need to better understand neuroinflammatory pain in females. This, paired with the fact that currently there is no long-term effective treatment for neuropathic pain furthers the need to evaluate how neuropathic pain develops in both sexes and how it can be relieved. Here we show that chronic constriction injury of the sciatic nerve caused comparable levels of mechanical allodynia in both sexes. Using a COX-2 inhibiting theranostic nanoemulsion with increased drug loading, both sexes achieved similar reduction in mechanical hypersensitivity. Given that both sexes have improved pain behavior, we specifically explored differential gene expression between sexes in the dorsal root ganglia (DRG) during pain and relief. Total RNA from the DRG revealed a sexually dimorphic expression for injury and relief caused by COX-2 inhibition. Of note, both males and females experience increased expression of activating transcription factor 3 (Atf3), however, only the female DRG shows decreased expression following drug treatment. Alternatively, S100A8 and S100A9 expression appear to play a sex specific role in relief in males. The sex differences in RNA expression reveal that comparable behavior does not necessitate the same gene expression.

Targeted cyclooxygenase-2 inhibiting nanomedicine results in pain-relief and differential expression of the RNA transcriptome in the dorsal root ganglia of injured male rats.

Chronic constriction injury of the sciatic nerve in rats causes peripheral neuropathy leading to pain-like behaviors commonly seen in humans. Neuropathy is a leading cause of neuropathic pain, which involves a complex cellular and molecular response in the peripheral nervous system with interactions between neurons, glia, and infiltrating immune cells. In this study, we utilize a nonsteroidal anti-inflammatory drug -loaded nanoemulsion to deliver the cyclooxygenase-2 inhibitor, Celecoxib, directly to circulating monocytes following nerve injury, which provides long-lasting pain relief. However, it is not fully understood how cyclooxygenase-2 inhibition in a macrophage traveling to the site of injury impacts gene expression in the dorsal root ganglia. To elucidate aspects of the molecular mechanisms underlying pain-like behavior in chronic constriction injury, as well as subsequent pain relief with treatment, we employ RNAseq transcriptome profiling of the dorsal root ganglia associated with the injured sciatic nerve in rats. Using high throughput RNA sequencing in this way provides insight into the molecular mechanisms involved in this neuroinflammatory response. We compare the transcriptome from the dorsal root ganglias of the following study groups: chronic constriction injury animals administered with cyclooxygenase-2 inhibiting celecoxib-loaded nanoemulsion, chronic constriction injury animals administered with vehicle treatment, a drug-free nanoemulsion, and a group of naïve, unoperated and untreated rats. The results show an extensive differential expression of 115 genes. Using the protein annotation through evolutionary relationship classification system, we have revealed pain-related signaling pathways and underlying biological mechanisms involved in the neuroinflammatory response. Quantitative polymerase chain reaction validation confirms expression changes for several genes. This study shows that by directly inhibiting cyclooxygenase-2 activity in infiltrating macrophages at the injured sciatic nerve, there is an associated change in the transcriptome in the cell bodies of the dorsal root ganglia.

Differential Expression of Neuroinflammatory mRNAs in the Rat Sciatic Nerve Following Chronic Constriction Injury and Pain-Relieving Nanoemulsion NSAID Delivery to Infiltrating Macrophages.

The neuroinflammatory response to peripheral nerve injury is associated with chronic pain and significant changes in the molecular expression profiles of mRNAs in neurons, glia and infiltrating immune cells. Chronic constriction injury (CCI) of the rat sciatic nerve provides an opportunity to mimic neuropathic injury and quantitatively assess behavior and differential gene expression in individual animals. Previously, we have shown that a single intravenous injection of nanoemulsion containing celecoxib (0.24 mg/kg) reduces inflammation of the sciatic nerve and relieves pain-like behavior for up to 6 days. Here, we use this targeted therapy to explore the impact on mRNA expression changes in both pain and pain-relieved states. Sciatic nerve tissue recovered from CCI animals is used to evaluate the mRNA expression profiles utilizing quantitative PCR. We observe mRNA changes consistent with the reduced recruitment of macrophages evident by a reduction in chemokine and cytokine expression. Furthermore, genes associated with adhesion of macrophages, as well as changes in the neuronal and glial mRNAs are observed. Moreover, genes associated with neuropathic pain including Maob, Grin2b/NMDAR2b, TrpV3, IL-6, Cacna1b/Cav2.2, Itgam/Cd11b, Scn9a/Nav1.7, and Tac1 were all found to respond to the celecoxib loaded nanoemulsion during pain relief as compared to those animals that received drug-free vehicle. These results demonstrate that by targeting macrophage production of PGE2 at the site of injury, pain relief includes partial reversal of the gene expression profiles associated with chronic pain.

Backward design as a mobile application development strategy.

Backward design is a well-established design strategy that has been used to produce educational curriculum for decades. While traditionally used to plan and create classroom-based curriculum, in this paper we explore the use of backward design as a design strategy for the development of an educational mobile application, BiblioTech™ "CityHacks: In Search of Sleep." We discuss the process from initial conception to launch and updates, as well as plans for future research.

Summer undergraduate research: A new pipeline for pain clinical practice and research.

BACKGROUND: Most medical schools fail to provide adequate training of clinicians in the treatment of pain. Similarly, despite the fact that over 1/3 of Americans suffer from chronic pain, National Institutes of Health (NIH) funding for pain represents only ~1% of the NIH budget. These issues may dissuade students from pursing pain in their clinical and research careers. To address these gaps in training and funding, we argue that exposing students to pain science early in their careers, at the undergraduate level, may be an effective method to develop a pipeline for future pain clinicians and scientists. To highlight our argument, we will describe our recent successful implementation of a cross-disciplinary and community-engaged biomedical summer research program. The Pain Undergraduate Research Experience (PURE) summer program involved both off-site and on-site experiences to expose undergraduate students to the range of careers in the pain field from basic science to clinical practice. The objective of the 10-week long PURE program was to evaluate whether a combination of basic science research, clinical practice visits, and patient interactions would increase student understanding of and exposure to the underlying science of pain. METHODS: A pre-post cohort study was used without a comparison group. Entry and exit surveys were used to evaluate students' perceptions about pain clinical practice and research, student interest in pain, and student confidence about communicating about pain and doing basic science pain research. RESULTS: Students reported significant increases to a number of questions in the survey. Questions were scored on 5 point Likert scales and there was significant increases in student understanding of what life is like with chronic pain (2.6 vs 4.3 post survey), their confidence in explaining pain to a patient (2.8 vs 4.1) or researcher (2.8 vs 4), and their comfort with pain terminology(2.8 vs 3.9). CONCLUSIONS: With the PURE program, we wanted to entice top undergraduates to consider pain as a future area of study, practice, and/or research. We present a model that can be easily implemented at research universities throughout the United States.

Folate-conjugated near-infrared fluorescent perfluorocarbon nanoemulsions as theranostics for activated macrophage COX-2 inhibition.

Activated macrophages play a critical role in the orchestration of inflammation and inflammatory pain in several chronic diseases. We present here the first perfluorocarbon nanoemulsion (PFC NE) that is designed to preferentially target activated macrophages and can deliver up to three payloads (two fluorescent dyes and a COX-2 inhibitor). Folate receptors are overexpressed on activated macrophages. Therefore, we introduced a folate-PEG-cholesterol conjugate into the formulation. The incorporation of folate conjugate did not require changes in processing parameters and did not change the droplet size or fluorescent properties of the PFC NE. The uptake of folate-conjugated PFC NE was higher in activated macrophages than in resting macrophages. Flow cytometry showed that the uptake of folate-conjugated PFC NE occurred by both phagocytosis and receptor-mediated endocytosis. Furthermore, folate-conjugated PFC NE inhibited the release of proinflammatory cytokines (TNF-α and IL-6) more effectively than nonmodified PFC NE, while drug loading and COX-2 inhibition were comparable. The PFC NEs reported here were successfully produced on multiple scales, from 25 to 200 mL, and by using two distinct processors (microfluidizers: M110S and LM20). Therefore, folate-conjugated PFC NEs are viable anti-inflammatory theranostic nanosystems for macrophage drug delivery and imaging.

The radiological management of pseudoaneurysms complicating pancreatitis.

CONTEXT: Pseudoaneurysms associated with pancreatitis are rare, and bleeding pseudoaneurysms are associated with a high mortality. OBJECTIVE: The aim of this study was to report the outcomes of endovascular and percutaneous therapy in the management of pseudoaneurysms secondary to pancreatitis. PATIENTS: Patients who underwent angiography for pseudoaneurysms associated with pancreatitis from 2005 to 2011 were identified from the angiography database. MAIN OUTCOME MEASURES: Patient demographics, clinical presentation, radiological findings, treatment, and outcomes were retrospectively reviewed. RESULTS: Nineteen pseudoaneurysms associated with pancreatitis in 13 patients were identified. The diagnosis of a pseudoaneurysm was made by computerised tomography angiography in seven patients, followed by portal venous phase contrast enhanced CT (n=4), duplex ultrasound (n=1) and angiography (n=1). At angiography, coil embolisation was attempted in 11 patients with an initial success rate of 82% (n=9). One patient underwent successful embolisation with percutaneous thrombin injection. The recurrence rate following initial successful embolisation was 11% (n=1). There were no episodes of re-bleeding following embolisation but re-bleeding following thrombin injection was observed in one case. The morbidity and mortality rate in the 12 patients that were successfully treated was 25% (n=3) and 8% (n=1), respectively. All 12 patients that were successfully treated demonstrated radiological resolution of their pseudoaneurysms, with a median follow-up of 20 months. CONCLUSION: Endovascular embolisation is a suitable first-line management strategy associated with low recurrence rates. The role of percutaneous thrombin injection is yet to be defined.

Behavioral and inflammatory sex differences revealed by celecoxib nanotherapeutic treatment of peripheral neuroinflammation.

Neuropathic pain affects millions of people worldwide, yet the molecular mechanisms of how it develops and persists are poorly understood. Given that males have historically been utilized as the primary sex in preclinical studies, less is known about the female neuroinflammatory response to injury, formation of pain, or response to pain-relieving therapies. Macrophages contribute to the development of neuroinflammatory pain via the activation of their cyclooxygenase-2 (COX-2) enzyme, which leads to the production of prostaglandin E2 (PGE2). PGE2 activates nociception and influences additional leukocyte infiltration. Attenuation of COX-2 activity decreases inflammatory pain, most commonly achieved by nonsteroidal anti-inflammatory drugs (NSAIDs), yet NSAIDs are considered ineffective for neuropathic pain due to off target toxicity. Using chronic constriction injury of the rat sciatic nerve, we show that males and females exhibit quantitatively the same degree of mechanical allodynia post injury. Furthermore, a low-dose nanotherapeutic containing the NSAID celecoxib is phagocytosed by circulating monocytes that then naturally accumulate at sites of injury as macrophages. Using this nanotherapeutic, we show that treated males exhibit complete reversal of hypersensitivity, while the same dose of nanotherapeutic in females provides an attenuated relief. The difference in behavioral response to the nanotherapy is reflected in the reduction of infiltrating macrophages at the site of injury. The observations contained in this study reinforce the notion that female neuroinflammation is different than males.

Real-time, spatial decision support to optimize malaria vector control: The case of indoor residual spraying on Bioko Island, Equatorial Guinea.

Public health interventions require evidence-based decision-making to maximize impact. Spatial decision support systems (SDSS) are designed to collect, store, process and analyze data to generate knowledge and inform decisions. This paper discusses how the use of a SDSS, the Campaign Information Management System (CIMS), to support malaria control operations on Bioko Island has impacted key process indicators of indoor residual spraying (IRS): coverage, operational efficiency and productivity. We used data from the last five annual IRS rounds (2017 to 2021) to estimate these indicators. IRS coverage was calculated as the percentage of houses sprayed per unit area, represented by 100x100 m map-sectors. Optimal coverage was defined as between 80% and 85%, and under and overspraying as coverage below 80% and above 85%, respectively. Operational efficiency was defined as the fraction of map-sectors that achieved optimal coverage. Daily productivity was expressed as the number of houses sprayed per sprayer per day (h/s/d). These indicators were compared across the five rounds. Overall IRS coverage (i.e. percent of total houses sprayed against the overall denominator by round) was highest in 2017 (80.2%), yet this round showed the largest proportion of oversprayed map-sectors (36.0%). Conversely, despite producing a lower overall coverage (77.5%), the 2021 round showed the highest operational efficiency (37.7%) and the lowest proportion of oversprayed map-sectors (18.7%). In 2021, higher operational efficiency was also accompanied by marginally higher productivity. Productivity ranged from 3.3 h/s/d in 2020 to 3.9 h/s/d in 2021 (median 3.6 h/s/d). Our findings showed that the novel approach to data collection and processing proposed by the CIMS has significantly improved the operational efficiency of IRS on Bioko. High spatial granularity during planning and deployment together with closer follow-up of field teams using real-time data supported more homogeneous delivery of optimal coverage while sustaining high productivity.

A family-centered educational program to promote independence in pediatric heart transplant recipients.

CONTEXT: Characteristic adolescent risk-taking behavior, including nonadherence with prescribed medications, can be life-threatening for transplant recipients. Suggestions for managing nonadherence in teen recipients include providing them and their parents with adequate information about medications, talking with and listening to pediatric recipients about problems with the comprehensive regimen, and encouraging age-appropriate responsibility for maintaining health. OBJECTIVE: The clinical goal of this project was to develop a structured age-appropriate educational program to prepare pediatric transplant recipients and their families for the patient's life as a responsible, independent individual. Our primary research goal was to assess patients' and parents' knowledge about critical aspects of heart transplantation and the treatment regimen with brief questionnaires before and after they received the educational materials from their primary nurse coordinator. DESIGN, SETTING, PARTICIPANTS: This descriptive pre-post test study was done to assess the effectiveness of an innovative family-centered educational program among 20 pediatric heart transplant recipients and their parents at Children's Hospital of Pittsburgh, University of Pittsburgh Medical Center. MAIN OUTCOME MEASURE, RESULTS: Percentage change in children's scores on questionnaires given before and after the educational intervention ranged from -8% to 300% (mean, 64.1%). Percentage change in scores from before to after for parents ranged from -19% to 53.8% (mean, 7.2%).

Social capital or vulnerability: Which has the stronger connection with selected U.S. health outcomes?

We tested associations between social capital or vulnerability and health outcome measures of adult obesity, adult diabetes, and life expectancy at the county level in the United States with data from 2015 to 2018. This ecological cross-sectional study utilized secondary data from four open access databases: The Geography of Social Capital (U.S. Congress, 2018), County Health Rankings (2018), CDC's Behavioral Risk Factor Surveillance System (BRFSS, 2018) and the Kaiser Family Foundation (KFF, 2015). Our dependent variables were adult obesity, adult diabetes, and life expectancy. We identified the highest and lowest states' prevalence for each of three health outcomes in each of the four U.S. regions-Northeast, South, Midwest, and West. Each dependent variable was assessed using a sample of 32 counties (N = 32). Data analysis consisted of bivariate and regression analysis. Our results showed that the most consistent measure of "vulnerability" linked significantly to all three health conditions studied was percent births to unmarried women (Obesity p < .001; Diabetes p = .049; Life Expectancy p = .019). The most consistent measure of "social capital" linked to all three health conditions was recreation establishments per 1,000 inhabitants (Obesity p = .006; Diabetes p = .005; Life Expectancy p = .018). We concluded that measures of vulnerability were strongly associated with obesity, diabetes, and life expectancy when compared with social capital indicators. However, measures of social capital consistently accounted for the second-greatest proportion of the variance. Social and community contexts should be constantly addressed by both public health governmental- and scholarly-research agendas in the United States.

Induction of parturition in a large number of pregnant dairy goats and its benefits as a management tool in a commercial scale goat operation.

At LFB USA, Inc., transgenic goats are utilized for the production of recombinant human protein therapeutics in their milk through the rPRO™ Technology platform. This retrospective analysis and report describes the results of induced parturition and its use as a management tool in this large herd of dairy goats. Over a three-year period, 342 does received pronuclear microinjected (MI) embryos transferred into the oviductal lumen via midline laparotomy (day 1). To initiate the induction process, does were given intramuscular injections (IM) of 10 mg each of prostaglandin (Lutalyse®) and dexamethasone to induce parturition on days 144-148 of pregnancy. Mean and Standard Deviation (±SD) time to parturition was 36.7 (±6.5) hours. Does were given these injections at 4pm on Sundays with an expected kidding time of late Monday into Tuesday morning. Of the 342 does, 333 or 97% had kidded by 3pm the following Tuesday, and 313 or 91% kidded in the 18 h between 9pm Monday and 3pm on Tuesday or between 29 and 47 h post induction. By the end of Tuesday, most kids had received colostrum and were transferred to the nursery. The incidences of kid mortality and retained placenta were 2.5% and 1.5%, respectively, clearly achieving a priority at this commercial operation for generating a high percentage of live kids (97.5%) of marked value being produced. The use of induced parturition allowed this large dairy operation to designate two 9-h time blocks in which to concentrate parturition times within the herd. This facilitated strategic scheduling to optimize availability of staff, in order to assist with parturition, separate kids from the dam at birth, and ensure adequate and prompt feeding of colostrum. Predicting the time of kidding in this way can serve as an effective management tool, especially to help reduce kid mortality and prevent disease spread by restricting suckling of colostrum.

How media empower the vulnerable: Using community structure theory to analyze relationships between demographics and health reporting.

Instead of studying the impact of media on society, the traditional "top down" orientation of most communication studies scholars, this keynote presentation adopted the opposite perspective, exploring the "bottom-up" impact of "society" on "media". Unlike conventional "agenda-setting theory", which suggests that nationally prominent news media set issue "agendas" for other news media and public opinion, and also unlike the "guard dog" view that media essentially protect the interests of political and economic elites, the "community structure theory" explores links between different community (typically city or nation-state) demographics and variations in reporting on critical health concerns. Summarizing his scholarship on health communication presented and published over decades, the speaker outlined community structure theory's illumination of two overall patterns in US and cross-national coverage of health communication issues. In US coverage, broad measures of economically "buffered" privilege (educational, income, or occupational advantage) are linked to "favorable" or "government responsibility" coverage of health issues, and specific measures of "health" privilege (physicians, hospitals) are connected to "favorable" or "government responsibility" coverage promoting selected health issues. In cross-national coverage, specific measures of national "health vulnerability" (such as percent without improved water access, infant mortality rate) are linked to "government" responsibility coverage for selected health issues (human trafficking, water handling/contamination). In addition, broad measures of "macro" vulnerability conditions (agricultural dependence, political instability) are associated with "government" responsibility coverage for a wide range of health issues (genetically modified foods, drug trafficking, condom promotion, and food security). Overall, community structure theory's "bottom up" perspective reveals how the vulnerable are empowered by their demographic alignment with variations in health communication.

The Use of a Mobile Application to Teach Concussion-Related Health Knowledge.

Sports-related concussions affect over 280,000 adolescents each year while the general public remains ill-informed about concussions, signs/symptoms, and treatments. Adolescents may be at an increased risk for experiencing adverse physiological and psychological effects from concussions, underscoring the critical need for effective concussion education strategies. While mobile apps are increasingly being used in education and healthcare settings, none were found to offer comprehensive concussion education capable of reaching diverse audiences. The interactive mobile app "Rebound: Beating Concussions" has the potential to be an effective teaching tool for school athletic programs and medical professionals to communicate important concussion-related information to student athletes, parents, and sports coaches. A mixed methods study was used to determine the app's ability to convey information about concussions to student athletes in grades 5 through 12, parents of student athletes, and sports coaches. Concussion knowledge and participant opinions were assessed via a pre/post model and administered before and after app use. Participants demonstrated knowledge gains in the identification of concussion symptoms, treatments, and misconceptions. Additionally, participants demonstrated positive opinions on the content of the app, its relevance to everyday life, and its potential as a teaching tool.

Ttk69-dependent repression of lozenge prevents the ectopic development of R7 cells in the Drosophila larval eye disc.

BACKGROUND: During the development of the Drosophila eye, specific cell types differentiate from an initially equipotent group of uncommitted precursor cells. The lozenge (lz) gene, which is a member of the Runt family of transcriptional regulators, plays a pivotal role in mediating this process through regulating the expression of several fate-specifying transcription factors. However, the regulation of lz, and the control of lz expression levels in different cell types is not fully understood. RESULTS: Here, we show a genetic interaction between Tramtrack69 (Ttk69) a key transcriptional repressor and an inhibitor of neuronal fate specification, and lz, the master patterning gene of cells posterior to the morphogenetic furrow in the Drosophila eye disc. Loss of Ttk69 expression causes the development of ectopic R7 cells in the third instar eye disc, with these cells being dependent upon Lz for their development. Using the binary UAS Gal4 system, we show that overexpression of Ttk69 causes the loss of lz-dependent differentiating cells, and a down-regulation of Lz expression in the developing eye. The loss of lz-dependent cells can be rescued by overexpressing lz via a GMR-lz transgene. We provide additional data showing that factors functioning upstream of Ttk69 in eye development regulate lz in a Ttk69-dependent manner. CONCLUSIONS: Our results lead us to conclude that Ttk69 can either directly or indirectly repress lz gene expression to prevent the premature development of R7 precursor cells in the developing eye of Drosophila. We therefore define a mechanism for the tight regulatory control of the master pre-patterning gene, lz, in early Drosophila eye development and provide insight into how differential levels of lz expression can be achieved to effect specific cell fate outcomes.

Nanomedicine-driven neuropathic pain relief in a rat model is associated with macrophage polarity and mast cell activation.

We explored the immune neuropathology underlying multi-day relief from neuropathic pain in a rat model initiated at the sciatic nerve, by using a nanoemulsion-based nanomedicine as a biological probe. The nanomedicine is theranostic: both therapeutic (containing celecoxib drug) and diagnostic (containing near-infrared fluorescent (NIRF) dye) and is small enough to be phagocytosed by circulating monocytes. We show that pain-like behavior reaches a plateau of maximum hypersensitivity 8 days post-surgery, and is the rationale for intravenous delivery at this time-point. Pain relief is evident within 24 h, lasting approximately 6 days. The ipsilateral sciatic nerve and associated L4 and L5 dorsal root ganglia (DRG) tissue of both nanomedicine and control (nanoemulsion without drug) treated animals was investigated by immunofluorescence and confocal microscopy at the peak of pain relief (day-12 post-surgery), and when pain-like hypersensitivity returns (day-18 post-surgery). At day-12, a significant reduction of infiltrating macrophages, mast cells and mast cell degranulation was observed at the sciatic nerve following treatment. In the DRG, there was no effect of treatment at both day-12 and day-18. Conversely, at the DRG, there is a significant increase in macrophage infiltration and mast cell degranulation at day-18. The treatment effect on immune pathology in the sciatic nerve was investigated further by assessing the expression of macrophage cyclooxygenase-2 (COX-2)-the drug target-and extracellular prostaglandin E2 (PGE2), as well as the proportion of M1 (pro-inflammatory) and M2 (anti-inflammatory) macrophages. At day-12, there is a significant reduction of COX-2 positive macrophages, extracellular PGE2, and a striking reversal of macrophage polarity. At day-18, these measures revert to levels observed in control-treated animals. Here we present a new paradigm of immune neuropathology research, by employing a nanomedicine to target a mechanism of neuropathic pain-resulting in long-lasting pain relief--whilst revealing novel immune pathology at the injured nerve and associated DRG.