Erythromycin Improves the Quality of Esophagogastroduodenoscopy in Upper Gastrointestinal Bleeding: A Network Meta-Analysis.

BACKGROUND/AIM: Upper gastrointestinal bleeding (UGIB) usually requires esophagogastroduodenoscopy (EGD) for diagnostic and-potentially-therapeutic purposes. However, blood within the gastric lumen may hinder the procedure. Administration of prokinetics like erythromycin has shown efficacy. This network meta-analysis investigates the efficacy of this intervention prior to EGD. METHODS: We performed a systematic literature search of Embase, PubMed/Medline, and other databases through March 8, 2022 to include randomized controlled trials (RCTs) comparing prokinetic use in EGD for UGIB. We used the DerSimonian-Laird approach to pool data and compare outcomes including need for repeat endoscopy and blood transfusion. Pooled prevalence of proportional outcomes, 95% confidence interval (CI), and p-values were calculated. RESULTS: We included eight RCTs with four distinct intervention groups (erythromycin, placebo to erythromycin, nasogastric (NG) lavage and NG lavage + erythromycin) published between 2002 and 2020 with a total of 721 patients (mean age 60.0 ± 3.1 years; 73.2% male). The need for second look endoscopy was significantly lower with erythromycin than placebo (relative risk: 0.42, CI 0.22-0.83, p = 0.01). Using the frequentist approach, the combination of NG lavage and erythromycin (92.2) was rated highest, followed by erythromycin alone (73.1) for higher rates of empty stomach. Erythromycin was rated highest for lower need for packed red blood cell transfusion (72.8) as well as mean endoscopy duration (66.0). CONCLUSION: Erythromycin improved visualization at EGD, reduced requirements for blood transfusion and repeat EGD, and shortened hospital stay. The combination of erythromycin and NG lavage showed reduced mortality.

Dual Antiplatelet Therapy Does Not Increase Bleeding Risk in Percutaneous Gastrostomy Tube Placement: Network Meta-Analysis.

BACKGROUND/OBJECTIVE: Gastrostomy tube (G tube) is a commonly performed procedure for nutritional support. Current guidelines recommend discontinuation of dual antiplatelet therapy (DAPT) prior to G tube placement to reduce bleeding risk. AIMS: We aim to compare bleeding risk in single, dual and no antiplatelet therapy during G tube placement. METHODS: We searched PubMed, Embase, Cochrane, and Web of Sciences to include comparative studies evaluating single antiplatelet (aspirin, clopidogrel), dual antiplatelet (DAPT, aspirin and clopidogrel), and no antiplatelet therapy. Direct as well as network meta-analyses comparing these arms were performed. Risk Differences (RD) with confidence intervals were calculated. RESULTS: 12 studies with 8471 patients were included. On direct meta-analysis, there was no significant difference noted between DAPT compared to Aspirin (RD 0.001 95% CI - 0.012 to 0.014, p = 0.87), Clopidogrel (RD 0.001 95% CI - 0.009 to 0.010, p = 0.92) or no antiplatelet group (RD 0.007 95% CI - 0.011 to 0.026, p = 0.44). Results were consistent on network meta-analysis and no difference was noted in bleeding rates when comparing DAPT with Aspirin (RD 0.001, 95% CI - 0.007 to 0.01, p = 0.76), Clopidogrel (RD 0.001, 95% CI - 0.01 to 0.011, p = 0.90) and no antiplatelet group (RD 0.002, 95% CI - 0.007 to 0.012, p = 0.62). CONCLUSION: There is no significant difference in bleeding risk between DAPT, single antiplatelet or no antiplatelet therapy on a population level. On an individual level, risk of ischemic events should be weighed against the risk of bleeding based on patient circumstances and risk profile. Our findings offer to provide additional data to make an informed decision between patients and physicians to make clinical decisions by assessing individual risks and benefits for optimal care of complex patients.

Increased stroke severity and mortality in patients with SARS-CoV-2 infection: An analysis from the N3C database.

BACKGROUND: Studies from early in the COVID-19 pandemic showed that patients with ischemic stroke and concurrent SARS-CoV-2 infection had increased stroke severity. We aimed to test the hypothesis that this association persisted throughout the first year of the pandemic and that a similar increase in stroke severity was present in patients with hemorrhagic stroke. METHODS: Using the National Institute of Health National COVID Cohort Collaborative (N3C) database, we identified a cohort of patients with stroke hospitalized in the United States between March 1, 2020 and February 28, 2021. We propensity score matched patients with concurrent stroke and SARS-COV-2 infection and available NIH Stroke Scale (NIHSS) scores to all other patients with stroke in a 1:3 ratio. Nearest neighbor matching with a caliper of 0.25 was used for most factors and exact matching was used for race/ethnicity and site. We modeled stroke severity as measured by admission NIHSS and the outcomes of death and length of stay. We also explored the temporal relationship between time of SARS-COV-2 diagnosis and incidence of stroke. RESULTS: Our query identified 43,295 patients hospitalized with ischemic stroke (5765 with SARS-COV-2, 37,530 without) and 18,107 patients hospitalized with hemorrhagic stroke (2114 with SARS-COV-2, 15,993 without). Analysis of our propensity matched cohort revealed that stroke patients with concurrent SARS-COV-2 had increased NIHSS (Ischemic stroke: IRR=1.43, 95% CI:1.33-1.52, p<0.001; hemorrhagic stroke: IRR=1.20, 95% CI:1.08-1.33, p<0.001), length of stay (Ischemic stroke: estimate = 1.48, 95% CI: 1.37, 1.61, p<0.001; hemorrhagic stroke: estimate = 1.25, 95% CI: 1.06, 1.47, p=0.007) and higher odds of death (Ischemic stroke: OR 2.19, 95% CI: 1.79-2.68, p<0.001; hemorrhagic stroke: OR 2.19, 95% CI: 1.79-2.68, p<0.001). We observed the highest incidence of stroke diagnosis on the same day as SARS-COV-2 diagnosis with a logarithmic decline in counts. CONCLUSION: This retrospective observational analysis suggests that stroke severity in patients with concurrent SARS-COV-2 was increased throughout the first year of the pandemic.

Stage at diagnosis and survival among adolescents and young adults with lymphomas following the Affordable Care Act implementation in California.

Adolescents and young adults (AYAs, 15-39 years) are the largest uninsured population in the Unites States, increasing the likelihood of late-stage cancer diagnosis and poor survival. We evaluated the associations between the Affordable Care Act (ACA), insurance coverage, stage at diagnosis and survival among AYAs with lymphoma. We used data from the California Cancer Registry linked to Medicaid enrollment files on AYAs diagnosed with a primary non-Hodgkin (NHL; n = 5959) or Hodgkin (n = 5378) lymphoma pre-ACA and in the early and full ACA eras. Health insurance was categorized as continuous Medicaid, discontinuous Medicaid, Medicaid enrollment at diagnosis/uninsurance, other public and private. We used multivariable regression models for statistical analyses. The proportion of AYAs uninsured/Medicaid enrolled at diagnosis decreased from 13.4% pre-ACA to 9.7% with full ACA implementation, while continuous Medicaid increased from 9.3% to 29.6% during this time (P < .001). After full ACA, AYAs with NHL were less likely to be diagnosed with Stage IV disease (adjusted odds ratio [aOR] = 0.84, 95% confidence interval [CI] = 0.73-0.97). AYAs with lymphoma were more likely to receive care at National Cancer Institute-Designated Cancer Centers (aOR = 1.42, 95% CI = 1.28-1.57) and had lower likelihood of death (adjusted hazard ratio = 0.54, 95% CI = 0.46-0.63) after full ACA. However, AYAs from the lowest socioeconomic neighborhoods, racial/ethnic minority groups and those with Medicaid continued to experience worse survival. In summary, AYAs with lymphomas experienced increased access to healthcare and better clinical outcomes following Medicaid expansion under the ACA. Yet, socioeconomic and racial/ethnic disparities remain, calling for additional efforts to decrease health inequities among underserved AYAs with lymphoma.

Environmental pesticide exposure and non-Hodgkin lymphoma survival: a population-based study.

BACKGROUND: There is evidence indicating that pesticide exposure is a risk factor for non-Hodgkin lymphoma (NHL) development. However, the association between pesticide exposure and NHL survival is not well-established. METHODS: Using the California Cancer Registry, we identified patients with a first primary diagnosis of NHL from 2010 to 2016 and linked these patients with CalEnviroScreen 3.0 to obtain production agriculture pesticide exposure to 70 chemicals from the state-mandated Pesticide Use Reporting (PUR) by census tract from 2012 to 2014. In addition, data from PUR was integrated into a geographic information system that employs land-use data to estimate cumulative exposure to specific pesticides previously associated with NHL (glyphosate, organophosphorus, carbamate, phenoxyherbicide, and 2,4-dimethylamine salt) between 10 years prior up to 1 year after NHL diagnosis. Multivariable Cox proportional hazards regression models were used to evaluate the association between total pesticide exposure from CalEnviroScreen 3.0 and individual pesticide exposure from geographic land use data and lymphoma-specific and overall survival. RESULTS: Among 35,808 NHL patients identified, 44.2% were exposed to pesticide in their census tract of residence. Glyphosate, organophosphorus, carbamate, phenoxyherbicide, and 2,4-dimethylamine salt exposure was observed in 34.1%, 26.0%, 10.6%, 14.0%, and 12.8% of NHL patients, respectively. Total pesticide exposure at the time of diagnosis was not associated with lymphoma-specific or overall survival. In addition, no association was consistently found between glyphosate, organophosphorus, carbamate, phenoxyherbicide, and 2,4 dimethylamine salt exposure and lymphoma-specific or overall survival. CONCLUSIONS: Although we found no consistent associations between agricultural pesticide exposure at the neighborhood level and worse survival, these results provide a platform for designing future studies to determine the association between pesticide and NHL.

IL-37 is increased in brains of children with autism spectrum disorder and inhibits human microglia stimulated by neurotensin.

Autism spectrum disorder (ASD) does not have a distinct pathogenesis or effective treatment. Increasing evidence supports the presence of immune dysfunction and inflammation in the brains of children with ASD. In this report, we present data that gene expression of the antiinflammatory cytokine IL-37, as well as of the proinflammatory cytokines IL-18 and TNF, is increased in the amygdala and dorsolateral prefrontal cortex of children with ASD as compared to non-ASD controls. Gene expression of IL-18R, which is a receptor for both IL-18 and IL-37, is also increased in the same brain areas of children with ASD. Interestingly, gene expression of the NTR3/sortilin receptor is reduced in the amygdala and dorsolateral prefrontal cortex. Pretreatment of cultured human microglia from normal adult brains with human recombinant IL-37 (1 to 100 ng/mL) inhibits neurotensin (NT)-stimulated secretion and gene expression of IL-1ß and CXCL8. Another key finding is that NT, as well as the proinflammatory cytokines IL-1ß and TNF increase IL-37 gene expression in cultured human microglia. The data presented here highlight the connection between inflammation and ASD, supporting the development of IL-37 as a potential therapeutic agent of ASD.

Symptomatic presentation of cervical cancer in emergency departments in California.

PURPOSE: Through screening and HPV vaccination, cervical cancer can mostly be prevented or detected very early, before symptoms develop. However, cervical cancer persists, and many women are diagnosed at advanced stages. Little is known about the degree to which U.S. women may begin their diagnostic workup for cervical cancer in Emergency Departments (ED). We sought to quantify the proportion of women presenting symptomatically in the ED prior to their diagnosis with cervical cancer and to describe their characteristics and outcomes. METHODS: We identified women diagnosed from 2006 to 2017 with cervical cancer in the California Cancer Registry. We linked this cohort to statewide ED discharge records to determine ED use and symptoms present at the encounter. Multivariable logistic regression models examined associations with ED use and multivariable Cox proportional hazards regression models examined associations with survival. RESULTS: Of the more than 16,000 women with cervical cancer in the study cohort, 28% presented symptomatically in the ED prior to diagnosis. Those presenting symptomatically were more likely to have public (odds ratio [OR] 1.16; 95% confidence interval [CI] 1.06-1.27) or no insurance (OR 4.81; CI 4.06-5.71) (vs. private), low socioeconomic status (SES) (OR 1.76; CI 1.52-2.04), late-stage disease (OR 5.29; CI 4.70-5.96), and had a 37% increased risk of death (CI 1.28-1.46). CONCLUSION: Nearly a third of women with cervical cancer presented symptomatically, outside of a primary care setting, suggesting that many women, especially those with low SES, may not be benefiting from screening or healthcare following abnormal results.

The influence of the Affordable Care Act-Dependent Care Expansion on insurance coverage among young cancer survivors in California: an updated analysis.

PURPOSE: To assess changes in health insurance coverage for young cancer patients pre- and post- the Affordable Care Act-Dependent Care Expansion (ACA-DCE) implementation in California. Further, we examined differences in insurance coverage by socioeconomic and race/ethnicity. METHODS: Data were obtained from the California Cancer Registry and Medicaid enrollment files, from 2005 to 2014. We conducted difference-in-difference analyses among 7042 cancer patients aged 22-25 years ("intervention group") and 25,269 aged 26-34 years ("control group"). We also examined the independent and combined effects of race/ethnicity and neighborhood socioeconomic status (nSES) on insurance coverage. RESULTS: After the ACA-DCE implementation, we observed a 52.7% reduction in the proportion of uninsured and a 35.7% increase in the proportion of privately insured patients. There was also a 17.3% reduction in Medicaid at cancer diagnosis and a 27.5% reduction in discontinuous Medicaid enrollment. However, these benefits were limited to patients of non-Hispanic White, Hispanic and Asian/Pacific Islander race/ethnicity living in higher nSES, with no differences in insurance enrollment among young adults who lived in low nSES or those of Black race/ethnicity. CONCLUSION: The ACA-DCE broadened insurance coverage for young adults with cancer in California. Yet, only certain subgroups of patients have benefited from this policy.

Impact of insurance type and timing of Medicaid enrollment on survival among adolescents and young adults with cancer.

BACKGROUND: Adolescents and young adults (AYAs) with public or no insurance experience later stage at diagnosis and worse overall survival compared with those with private insurance. However, prior studies have not distinguished the survival impact of continuous Medicaid coverage prior to diagnosis compared with gaining Medicaid coverage at diagnosis. METHODS: We linked a cohort of AYAs aged 15-39 who were diagnosed with 13 common cancers from 2005 to 2014 in the California Cancer Registry with California Medicaid enrollment files to ascertain Medicaid enrollment, with other insurance determined from registry data. We used Cox proportional hazards regression to evaluate the impact of insurance on survival, adjusting for clinical and demographic characteristics. RESULTS: Among 62 218 AYAs, over 65% had private/military insurance, 10% received Medicaid at diagnosis, 13.2% had continuous Medicaid, 4.1% had discontinuous Medicaid, 1.7% had other public insurance, 3% were uninsured, and 2.6% had unknown insurance. Compared with those with private/military insurance, individuals with Medicaid insurance had significantly worse survival regardless of when coverage began (received Medicaid at diagnosis: hazard ratio [95% confidence interval]: 1.51 [1.42-1.61]; continuously Medicaid insured: 1.42 [1.33-1.52]; discontinuous Medicaid: 1.64 [1.49, 1.80]). Analyses of those with Medicaid insurance only demonstrated slightly worse cancer-specific survival among those with discontinuous Medicaid or enrollment at diagnosis compared with those with continuous enrollment, but results were not significant stratified by cancer site. CONCLUSIONS AND RELEVANCE: AYAs with Medicaid insurance experience worse cancer-specific survival compared with those with private/military insurance, yet continuous enrollment demonstrates slight survival improvements, providing potential opportunities for future policy intervention.

Adoption of Blockchain in Health Care.

This study aims to review current issues regarding the application of blockchain technology in health care. We illustrated the various ways in which blockchain can solve current health care issues in three main arenas: data exchange, contracts, and supply chain management. This paper presents several current and projected uses of blockchain technology in the health care industry. We predicted which of these applications are likely to be adopted quickly and provided a supply chain example of tracking the transportation of organs for transplantation.

Neurotensin stimulates sortilin and mTOR in human microglia inhibitable by methoxyluteolin, a potential therapeutic target for autism.

We had reported elevated serum levels of the peptide neurotensin (NT) in children with autism spectrum disorders (ASD). Here, we show that NT stimulates primary human microglia, the resident immune cells of the brain, and the immortalized cell line of human microglia-SV40. NT (10 nM) increases the gene expression and release (P < 0.001) of the proinflammatory cytokine IL-1ß and chemokine (C-X-C motif) ligand 8 (CXCL8), chemokine (C-C motif) ligand 2 (CCL2), and CCL5 from human microglia. NT also stimulates proliferation (P < 0.05) of microglia-SV40. Microglia express only the receptor 3 (NTR3)/sortilin and not the NTR1 or NTR2. The use of siRNA to target sortilin reduces (P < 0.001) the NT-stimulated cytokine and chemokine gene expression and release from human microglia. Stimulation with NT (10 nM) increases the gene expression of sortilin (P < 0.0001) and causes the receptor to be translocated from the cytoplasm to the cell surface, and to be secreted extracellularly. Our findings also show increased levels of sortilin (P < 0.0001) in the serum from children with ASD (n = 36), compared with healthy controls (n = 20). NT stimulation of microglia-SV40 causes activation of the mammalian target of rapamycin (mTOR) signaling kinase, as shown by phosphorylation of its substrates and inhibition of these responses by drugs that prevent mTOR activation. NT-stimulated responses are inhibited by the flavonoid methoxyluteolin (0.1-1 µM). The data provide a link between sortilin and the pathological findings of microglia and inflammation of the brain in ASD. Thus, inhibition of this pathway using methoxyluteolin could provide an effective treatment of ASD.

TNF stimulates IL-6, CXCL8 and VEGF secretion from human keratinocytes via activation of mTOR, inhibited by tetramethoxyluteolin.

Psoriasis is an autoimmune skin disease characterized by keratinocyte hyperproliferation and chronic inflammation. The pathogenesis of psoriasis involves proinflammatory cytokines, such as tumor necrosis factor (TNF), but the mechanism of keratinocyte activation is not well understood. Here, we show that TNF (10 or 50 ng/mL) stimulates a significant (P < .0001) gene expression and secretion of proinflammatory IL-6, CXCL8 and VEGF from both cultured human HaCaT and normal epidermal human keratinocytes (NHEKs). This effect occurs via activation of the mammalian target of rapamycin (mTOR) signalling complex as shown by Western blot analysis and phospho-ELISAs. Pretreatment with the novel natural flavonoid tetramethoxyluteolin (10-100 µmol L-1 ) significantly (P < .0001) inhibits gene expression and secretion (P < .0001) of all 3 mediators in a concentration-dependent manner. Moreover, tetramethoxyluteolin (50 µmol L-1 ) appears to be a potent inhibitor of the phosphorylated mTOR substrates (pmTORSer2448 , pp70S6KThr389 and p4EBP1Thr37/46 ) as compared to known mTOR inhibitors in keratinocytes. The present findings indicate that TNF stimulates skin inflammation via mTOR signalling. Inhibition by tetramethoxyluteolin may be used in the treatment for psoriasis.

Methoxyluteolin Inhibits Neuropeptide-stimulated Proinflammatory Mediator Release via mTOR Activation from Human Mast Cells.

Mast cells (MCs) are critical for allergic reactions but are also important in inflammatory processes. Stimulation by neuropeptides, such as substance P (SP) and neurotensin (NT), leads to release of preformed molecules stored in numerous MC secretory granules and newly synthesized proinflammatory mediators, including tumor necrosis factor, C-X-C motif chemokine ligand 8, and vascular endothelial growth factor. Here, we investigate the role of mammalian target of rapamycin (mTOR) signaling in the stimulation of cultured human LAD2 MCs by NT or SP, as well as the inhibitory effect of the natural flavonoids 3',4',5,7-tetrahydroxyflavone (luteolin) and its novel structural analog 3',4',5,7-tetramethoxyflavone (methoxyluteolin). Stimulation by NT (10 µM) or SP (1 µM) increases (P < 0.0001) gene expression (after 6 hours) and release (after 24 hours) of tumor necrosis factor, C-X-C motif chemokine ligand 8, and vascular endothelial growth factor. This occurs via activation of both mTOR complexes, as denoted by the increased phosphorylated (p) protein levels (P < 0.0001) of the downstream mTORC1 substrate pp70S6KThr389 and mTORC2 component pmTORSer2448. Pretreatment of human MCs using the mTORC1 inhibitor rapamycin, the mTORC1/mTORC2 inhibitor Torin1, or the two flavonoids decreases both gene expression and release (P < 0.0001) of all three mediators. Methoxyluteolin is a more potent human MC inhibitor than luteolin or Torin1, implicating other MC protein targets in addition to the mTOR complex. These findings indicate that mTOR is partially involved in the neuropeptide stimulation of MCs, but the novel flavonoid methoxyluteolin inhibits the response entirely, suggesting that it may be developed for treatment of allergic and inflammatory diseases.

The novel flavone tetramethoxyluteolin is a potent inhibitor of human mast cells.

BACKGROUND: Mast cells (MCs) are hematopoietic cells that mature in tissues and are involved in allergy, immunity, and inflammation by secreting multiple mediators. The natural flavone luteolin has anti-inflammatory actions and inhibits human mast cells (MCs). OBJECTIVE: We sought to investigate the ability of luteolin and its novel structural analog 3',4',5,7-tetramethoxyluteolin (methlut) to inhibit human MC mediator expression and release in vitro and in vivo. METHODS: Human LAD2 cells and umbilical primary human cord blood-derived cultured mast cells were stimulated with substance P (SP) or IgE/anti-IgE with or without preincubation with luteolin, methlut, or cromolyn (1-100 µmol/L) for 2 or 24 hours, after which mediator secretion was measured. The effect of the compounds on MC intracellular calcium levels and nuclear factor κB activation was also investigated. Pretreatment with methlut was also studied in mice passively sensitized with dinitrophenol-human serum albumin and challenged intradermally. RESULTS: Methlut is a more potent inhibitor than luteolin or cromolyn for ß-hexosaminidase and histamine secretion from LAD2 cells stimulated by either SP or IgE/anti-IgE, but only methlut and luteolin significantly inhibit preformed TNF secretion. Methlut is also a more potent inhibitor than luteolin of de novo-synthesized TNF from LAD2 cells and of CCL2 from human cord blood-derived cultured MCs. This mechanism of action for methlut might be due to its ability to inhibit intracellular calcium level increases, as well as nuclear factor κB induction, at both the transcriptional and translational levels in LAD2 cells stimulated by SP without affecting cell viability. Intraperitoneal treatment with methlut significantly decreases skin vascular permeability of Evans blue dye in mice passively sensitized to dinitrophenol-human serum albumin and challenged intradermally. CONCLUSION: Methlut is a promising MC inhibitor for the treatment of allergic and inflammatory conditions.

Recording inpatient weight: implications for medicines administration.

AIM: To identify the proportions of hospital inpatients with recorded weights: among all patients, and among those receiving weight-dosed drug therapy. METHOD: Survey of clinical notes of hospital inpatients across a convenience sample of 11 secondary and tertiary referral hospitals in England and Wales in November 2011. RESULTS: 1068 patients were included, and 1061 patient clinical notes were available (99.3%). Nearly all paediatric patients had recorded weights (77/78; 98.7%). Half of adult inpatients had recorded weights (503/983, 51.2%). The proportion of adult inpatients with recorded weights varied by hospital, ranging from 13.5% to 92.5% (p<0.0001). In those receiving gentamicin or therapeutic-dose low molecular weight heparin (t-LMWH), only 64.5% (71/110) had a recorded weight. CONCLUSIONS: Half of adult inpatients, and two-thirds of those receiving gentamicin or t-LMWH, had recorded weights. There was significant variation in rates of weighing adult inpatients across hospitals. This may put patients at increased risk of side effects and problems resulting from malnutrition.

Apolipoprotein A-I binding to anionic vesicles and lipopolysaccharides: role for lysine residues in antimicrobial properties.

Human apolipoprotein A-I (apoA-I) is a 28kDa protein and a major component of high-density lipoproteins, mediating several essential metabolic functions related to heart disease. In the present study the potential protective role against bacterial pathogens was explored. ApoA-I suppressed bacterial growth of Escherichia coli and Klebsiella pneumoniae. The protein was able to bind lipopolysaccharides and showed a strong preference for bilayer vesicles made of phosphatidylglycerol over phosphatidylcholine. Lysine side chains of apoA-I were acetylated to evaluate the importance of electrostatic forces in the binding interaction with both membrane components. Electrophoresis properties, dot blot analysis, circular dichroism, and fluorescence spectroscopy to probe for changes in protein structure indicated that the acetylated protein displayed a strongly reduced lipopolysaccharide and phosphatidylglycerol binding. A mutant containing only the N-terminal domain of apoA-I also showed a reduced ability to interact with the membrane components, although to a lesser extent. These results indicate the potential for apoA-I to function as an antimicrobial protein and exerts this function through lysine residues.

A pyrene based fluorescence approach to study conformation of apolipoprotein E3 in macrophage-generated nascent high density lipoprotein.

Apolipoprotein E3 (apoE3) is an anti-atherogenic apolipoprotein with the ability to exist in lipid-free and lipoprotein-associated states. During atherosclerosis, its function in promoting cholesterol efflux from macrophages via the ATP-binding cassette transporter A1 (ABCA1) takes a prominent role, leading to generation of nascent high density lipoprotein (nHDL) particles. The objective of this study is to understand the conformation adopted by apoE3 in macrophage-generated nHDL using a fluorescence spectroscopic approach involving pyrene. Pyrene-labeled recombinant human apoE3 displayed a robust ability to stimulate ABCA1-mediated cholesterol efflux from cholesterol-loaded J774 macrophages (which do not express apoE), comparable to that elicited by unlabeled apoE3. The nHDL recovered from the conditioned medium revealed the presence of apoE3 by immunoblot analysis. A heterogeneous population of nHDL bearing exogenously added apoE3 was generated with particle size varying from ∼12 to ∼19 nm in diameter, corresponding to molecular mass of ∼450 to ∼700 kDa. The lipid: apoE3 ratio varied from ∼60:1 to 10:1. A significant extent of pyrene excimer emission was noted in nHDL, indicative of spatial proximity between Cys112 on neighboring apoE3 molecules similar to that noted in reconstituted HDL. Cross-linking analysis using Cys-specific cross-linkers revealed the predominant presence of dimers. Taken together the data indicate a double belt arrangement of apoE molecules on nHDL. A similar organization of the C-terminal tail of apoE on nHDL was noted when pyrene-apoEA277C(201-299) was used as the cholesterol acceptor. These studies open up the possibility of using exogenously labeled apoE3 to generate nHDL for structural and conformational analysis.

Copper nitroprusside analogue nanoparticles against melanoma: detailed in vitro and in vivo investigation.

Melanoma is the most invasive and lethal form of skin cancer that arises from the malignant transformation of specialized pigment-producing cell melanocytes. Nanomedicine represents an important prospect to mitigate the difficulties and provide significant benefits to cure melanoma. In the present study, we investigated in vitro and in vivo therapeutic efficacies of copper nitroprusside analogue nanoparticles (abbreviated as CuNPANP) towards melanoma. Initially, in vitro anti-cancer activities of CuNPANP towards melanoma cells (B16F10) were evaluated by several experiments such as [methyl-3H]-thymidine incorporation assay, cell cycle and apoptosis assays using FACS analysis, ROS generation using DCFDA, DHE and DAF2A reagents, internalization of nanoparticles through ICP-OES analysis, co-localization of the nanoparticles using confocal microscopy, JC-1 staining to investigate the mitochondrial membrane potential (MMP) and immunofluorescence studies to analyze the expressions of cytochrome-c, Ki-67, E-cadherin as well as phalloidin staining to analyze the cytoskeletal integrity. Further, the in vivo therapeutic effectiveness of the nanoparticles was established towards malignant melanoma by inoculating B16F10 cells in the dorsal right abdomen of C57BL/6J mice. The intraperitoneal administration of CuNPANP inhibited tumor growth and increased the survivability of melanoma mice. The in vivo immunofluorescence studies (Ki-67, CD-31, and E-cadherin) and TUNEL assay further support the anti-cancer and apoptosis-inducing potential of CuNPANP, respectively. Finally, various signaling pathways and molecular mechanisms involved in anti-cancer activities were further evaluated by Western blot analysis. The results altogether indicated the potential use of copper-based nanomedicines for the treatment of malignant melanoma.

Retention of α-helical structure by HDL mimetic peptide ATI-5261 upon extensive dilution represents an important determinant for stimulating ABCA1 cholesterol efflux with high efficiency.

ATI-5261 is a novel, single-helix peptide that stimulates cellular cholesterol efflux with high potency similar to native apolipoproteins on a molar basis. Presently we investigated structural features of the peptide that conferred cholesterol efflux activity. Analogs of ATI-5261 with amino acids arranged in reverse order or with individual arginine (R) to glutamine (Q) substitutions (i.e. R3Q, R14Q, or R23Q) stimulated ABCA1 dependent cholesterol efflux similar to ATI-5261. Consequently, neither the presence of specific positively charged residues nor their specific arrangement along the length of the peptide was necessary for mediating cholesterol efflux. Similarly, peptides composed of all d-amino acids stimulated cholesterol efflux efficiently, indicating a stereospecific component was not required for promotion of cholesterol efflux from macrophages. Removal of two or more positively charged residues (R3, 14→Q and R3, 14, 23→Q) however, greatly reduced the ability of ATI-5261 to mediate cellular cholesterol efflux. This was accompanied by a loss of α-helical structure upon dilution, indicating the secondary structure of individual peptide strands was important for stimulating cholesterol efflux. Surprisingly, peptides with removal of two or more positively charged residues retained the ability to bind phospholipid and adopt an α-helical structure. These data indicate that the propensity of a hydrophobic peptide to form an amphipathic α-helix is not sufficient to mediate cellular cholesterol efflux. Efficient stimulation of cholesterol efflux requires that ATI-5261 retain α-helical structure upon dilution.

Focal brain inflammation and autism.

Increasing evidence indicates that brain inflammation is involved in the pathogenesis of neuropsychiatric diseases. Autism spectrum disorders (ASD) are characterized by social and learning disabilities that affect as many as 1/80 children in the USA. There is still no definitive pathogenesis or reliable biomarkers for ASD, thus significantly curtailing the development of effective therapies. Many children with ASD regress at about age 3 years, often after a specific event such as reaction to vaccination, infection, stress or trauma implying some epigenetic triggers, and may constitute a distinct phenotype. ASD children respond disproportionally to stress and are also affected by food and skin allergies. Corticotropin-releasing hormone (CRH) is secreted under stress and together with neurotensin (NT) stimulates mast cells and microglia resulting in focal brain inflammation and neurotoxicity. NT is significantly increased in serum of ASD children along with mitochondrial DNA (mtDNA). NT stimulates mast cell secretion of mtDNA that is misconstrued as an innate pathogen triggering an auto-inflammatory response. The phosphatase and tensin homolog (PTEN) gene mutation, associated with the higher risk of ASD, which leads to hyper-active mammalian target of rapamycin (mTOR) signalling that is crucial for cellular homeostasis. CRH, NT and environmental triggers could hyperstimulate the already activated mTOR, as well as stimulate mast cell and microglia activation and proliferation. The natural flavonoid luteolin inhibits mTOR, mast cells and microglia and could have a significant benefit in ASD.