Exosomes in Allergic Airway Diseases.

PURPOSE OF REVIEW: This review will cover what is known regarding exosomes and allergy, and furthermore discuss novel mechanism of exosome-mediated immune modulation and metabolic regulation via the transfer of mitochondria. RECENT FINDINGS: Exosomes are nano-sized extracellular vesicles (EVs) derived from the endosome that play a direct role in governing physiological and pathological conditions by transferring bioactive cargo such as proteins, enzymes, nucleic acids (miRNA, mRNA, DNA), and metabolites. Recent evidence suggest that exosomes may signal in autocrine but, most importantly, in paracrine and endocrine manner, being taken up by neighboring cells or carried to distant sites. Exosomes also mediate immunogenic responses, such as antigen presentation and inflammation. In asthma and allergy, exosomes facilitate cross-talk between immune and epithelial cells, and drive site-specific inflammation through the generation of pro-inflammatory mediators like leukotrienes. Recent studies suggest that myeloid cell-generated exosomes transfer mitochondria to lymphocytes. Exosomes are nano-sized mediators of the immune system which can modulate responses through antigen presentation, and the transfer of pro- and anti-inflammatory mediators. In addition to conventional mechanisms of immune modulation, exosomes may act as a novel courier of functional mitochondria that is capable of modulating the recipient cells bioenergetics, resulting in altered cellular responses. The transfer of mitochondria and modulation of bioenergetics may result in immune activation or dampening depending on the context.

Exosomes in immunoregulation of chronic lung diseases.

Exosomes are nano-sized, membrane-bound vesicles released from cells that transport cargo including DNA, RNA, and proteins, between cells as a form of intercellular communication. In addition to their role in intercellular communication, exosomes are beginning to be appreciated as agents of immunoregulation that can modulate antigen presentation, immune activation, suppression, and surveillance. This article summarizes how these multifaceted functions of exosomes may promote development and/or progression of chronic inflammatory lung diseases including asthma, chronic obstructive pulmonary disease, and pulmonary fibrosis. The potential of exosomes as a novel therapeutic is also discussed.

Injury in rugby league.

It was the purpose of this review to document the range, incidence, location and mechanism of injury occurring in the sport of rugby league. Rugby league is a collision sport played in Europe and the Pacific regions including Australia. The sport is well established and has competitions ranging from junior to elite professional. Due to the contact nature of the game, injury is relatively common. The most common injuries are musculotendinous in nature and afflict the lower limb more frequently than elsewhere. Despite the high incidence of minor (sprains/strains) to moderate musculoskeletal injury (fracture, ligament and joint injury) and minor head injuries such as lacerations, nasal fractures and concussions, rare more serious spinal cord and other injuries causing death have also been recorded. The literature on rugby league injury is small but growing and suffers from a lack of consistent definition of what an injury is, thereby causing variability in the nature and incidence/prevalence of injury. Information is lacking on the injury profiles of different age groups. Importantly, there has been little attempt to establish a coordinated injury surveillance program in rugby league in the junior or professional levels. The implementation of such programs would require a universal definition of injury and a focus on important events and competitions. The implementation could provide important information in the identification and prevention of risk factors for injury.

Synthesis and biological evaluation of new beta,beta'-disubstituted 6,7,8,9-tetrahydropyrido[1,2-a]indol-10-yl ethylamido melatoninergic ligands.

Tricyclic analogs of melatonin with alkyl and cycloalkyl moieties in the beta position of the ethylamido chain have been prepared and tested for their ability to activate pigment granule aggregation in Xenopus laevis melanophores. The introduction of two methyl groups in the beta position of the side-chain of the methoxyl-substituted ligands induces a synergistic effect in agonist potency, which, importantly, is maintained after the methoxyl substituent is removed. The presence of more bulky beta-substituents, regardless of the size of the R group, seems to lead to antagonism.

Polymorphisms of GSTP1 and related genes and prostate cancer risk.

Glutathione S-transferase P1 (GSTP1) is markedly downregulated in prostate cancer and prostatic intraepithelial neoplasia compared to normal prostate tissue. Downregulation of GSTP1 may, therefore, be an early event in prostate carcinogenesis. An A-->G polymorphism at nucleotide 313 results in an amino acid substitution (Ile105Val) in the substrate binding site of GSTP1 and reduces catalytic activity of GSTP1. In a study of 36 prostate cancer patients, Harries et al. reported that the Ile/Ile genotype is associated with a decreased risk of prostate cancer (odds ratio 0.4 (0.17-0.82)). We sought to confirm this finding and to examine the impact of this polymorphism together with several related polymorphisms implicated as risk factors for carcinogen-associated malignancies. One hundred and seventeen patients with prostate adenocarcinoma and 183 population-based controls were recruited to this case-control study. Genotyping of the GSTP1 (Ile105Val), GSTM1 (null), GSTT1 (null) and CYP1A1 (Ile462Val) genes was performed using polymerase chain reaction (PCR) based techniques on DNA prepared from peripheral blood. A questionnaire was used to collect demographic information from each subject. Cases were significantly older (P<0.0001) and had significantly greater family history of prostate cancer (P<0.0001), confirming known risk factors for this disease. By chi(2) analysis, none of the genotype distributions varied among cases and controls. Using a logistic regression model to control for known risk factors we were also unable to demonstrate a significant association with prostate cancer for any of the polymorphisms tested. This population fails to identify a relationship between the above polymorphisms and prostate adenocarcinoma.

Association of codon 72 polymorphism of p53 with lower prostate cancer risk.

BACKGROUND: A common germline polymorphism of p53 produces a protein with an Arg to Pro change at codon 72. This Pro variant has altered biochemical properties suggesting altered cancer susceptibility. METHODS: A case control study with 115 men with prostate cancer and 181 community control male subjects was conducted. Demographics, family history of cancer, and blood were obtained. Codon 72 genotypes were determined using PCR. RESULTS: The Pro/Pro genotype was associated with a markedly lower risk of prostate cancer (OR = 0.23, CI = 0.07-0.79, P = 0.012). Similar reduction in risk was observed when the analysis was limited to Caucasian subjects (86% of total). Reduction in risk remained significant in a logistic regression model after correcting for age and family history of prostate cancer (OR = 0.14, CI = 0.03-0.71, P = 0.017). CONCLUSIONS: Men with the p53 codon 72 Pro/Pro genotype appear to be at reduced risk of prostate cancer.

Weekly high-dose calcitriol and docetaxel in advanced prostate cancer.

Novel treatment regimens for androgen-independent prostate cancer (AIPC) are needed because currently available approaches have not been shown to improve survival. Docetaxel provides a good foundation for new therapeutic combinations because of its promising single-agent activity against prostate cancer and its favorable tolerability profile, particularly when administered weekly. In both tissue culture and animal models of prostate cancer, calcitriol (the biologically active form of vitamin D) enhanced the activity of docetaxel, paclitaxel, and platinum compounds. These effects were particularly notable at supraphysiologic calcitriol concentrations. Weekly calcitriol dosing is associated with minimal toxicity and permits substantial dose escalation over the daily schedule. A weekly calcitriol dose of 0.5 microg/kg produces plasma calcitriol levels 25-fold higher than the physiologic range. In a preclinical study at the Oregon Health Sciences University, calcitriol 5 micromol/L plus docetaxel 0.15 nmol/L was at least additive in inhibiting PC-3 colony formation. A phase II study is evaluating weekly administration of 0.5 microg/kg calcitriol orally on day 1 followed by 36 mg/m(2) docetaxel intravenously on day 2 in patients with AIPC (repeated for 6 consecutive weeks of each 8-week cycle). At the time of a preliminary analysis, 11 patients had been enrolled and were actively being treated. All 5 patients who had completed 8 weeks of calcitriol/docetaxel treatment achieved prostate-specific antigen (PSA) reductions of > or =50%. Two of these patients had confirmatory assessments, both meeting the formal PSA response criteria. Treatment has been well tolerated, with 1 patient experiencing a self-limited grade 3 toxicity and no patients experiencing grade 4 or 5 toxicities.

Interactive effects of characteristics of defendant and mock juror on U.S. participants' judgment and sentencing recommendations.

The authors examined the effects of interactions (a) between defendant attractiveness and juror gender and (b) between defendant race and juror race on judgment and sentencing among 207 Black, Hispanic, and White participants in the United States. After reading a vehicular-homicide vignette in which the defendant's attractiveness and race varied, the participants rated guilt and recommended sentences. The women treated the unattractive female defendant more harshly than they treated the attractive female defendant; the men showed an opposite tendency. The Black participants showed greater leniency when the defendant was described as Black rather than White. The Hispanic participants showed an opposite trend, and the White participants showed no race-based leniency. The findings on racial effects were consistent (a) with in-group favorability bias among the Black participants and (b) with attribution effects unrelated to race among the White participants.

Synergistic effects of feeding and dexamethasone on serum leptin levels.

The objectives of this study were to determine the time course of the stimulatory effect of dexamethasone on serum leptin and whether it depends on food intake. Dexamethasone (4mg) was administered I.V. over 1 minute to healthy human volunteers (n=8) under fasting and feeding conditions (2000 kcal given at three meals over 7 hours). At 10 hours, serum leptin levels were increased only in the fed subjects (delta leptin 10.6+/-1.6 vs -2.4+/-1.9 ng/ml, p=0.01, n=8). To assess the interactive effect of food and dexamethasone on serum leptin, a subgroup (n=4) was studied under 4 conditions: 1) dexamethasone/fast; 2) dexamethasone/food; 3) saline/fast; 4) saline/food. Serum leptin declined from baseline under the fasting conditions, with or without dexamethasone. Feeding prevented the drop in serum leptin. In the dexamethasone/food condition, leptin levels rose from baseline after 7 hours and doubled after 10 hours (p<0.05). The rise in serum leptin was significantly greater in the food/dexamethasone condition compared to all other conditions (p<0.05). In summary, dexamethasone has no independent effect on serum leptin in the absence of food intake. Rather, dexamethasone appears to potentiate the food-induced increase in serum leptin. This synergism may be mediated by insulin and/or other factors associated with food ingestion.

Children's attitudes toward violence on television.

Children's attitudes toward television violence were studied. A 47-item questionnaire collecting attitudinal and personal information was administered to 316 children aged 11 to 16 years. Cluster analysis was used to split the participants into two groups based on their attitudes toward television violence. A stepwise discriminant function analysis was performed to determine which personal characteristics would predict group membership. The only significant predictor of attitudes toward violence on television was the amount of television watched on school days (p < .05), but we also found that the impact of other predictor variables may have been mediated by this factor.

Toxicity and immunologic effects of continuous infusion of recombinant human interleukin-2 administered by selective hepatic perfusion in dogs.

A preclinical pilot study was done to evaluate the effects of a continuous regional hepatic arterial infusion of human recombinant interleukin-2 (IL-2) in dogs with an infusion pump. Preliminary studies demonstrated the ability to culture canine lymphokine-activated killer (LAK) cells in vitro and a canine LAK cell 15Cr assay was developed with a canine tumor cell line (CTAC) with appropriate controls. An in vitro study of the stability of IL-2 in the pump was done with a bioassay and enzyme-linked immunosorbent assay for IL-2 that demonstrated the stability of IL-2 during a 14-day period at 37 degrees C. Infusions of 300, 600, and 1200 units/kg/hr IL-2 were tested in vivo in dogs. LAK cell and natural killer cell activity, blood counts, and hepatic and renal function were monitored for 1 month. No significant natural killer or LAK response or toxicity was found at the 300 unit/kg/hr level. Infusion of 600 units/kg/hr was associated with a significant increase of the cytotoxic activity of peripheral blood lymphocytes after 3 weeks of treatment. At the 1200 unit/kg/hr level, increased activity occurred at 1 week and thereafter. The only significant toxicity was a 15% increase in body weight occurring during the infusion of 1200 units/kg/hr. Results of renal and hepatic function studies remained normal except for a slight elevation of transaminase levels after 4 weeks of 1200 units/kg/hr. A significant rise in eosinophil count was noted at each dosage level. Results of autopsies were unremarkable. These data demonstrate that continuous hepatic arterial regional infusion with relatively low doses of IL-2 is able to stimulate a sustained in vivo peripheral blood LAK cell effect in dogs with the absence of major side effects. These findings suggest that these methods may have both research application in large animals and clinical application in patients with tumors that are responsive to LAK cell lysis.

The nude mouse as a model for the study of human pancreatic cancer.

The purpose of this study was to characterize an in vivo model of human pancreatic cancer suitable for chemotherapy and immunotherapy studies. In this study we report a 2-year experience in growing the MIA PaCa-2 (CRL 1420) human pancreatic cancer cell line in 92 adult (8 weeks old) and 256 young (3-6 weeks old) nude mice. Ten million tumor cells were transplanted into orthotopic (duodenal lobe of the pancreas) and/or heterotopic positions (hepatic and subcutaneous) and data on operative mortality, effect of total body irradiation (TBI), tumor growth kinetics, and survival are presented comparing the two age groups. Operative mortality was due to anesthetic intolerance which was higher in the young mouse population (13.4% versus 5.7%). Adult mice withstood TBI (500 rad) without mortality but young mice were highly sensitive to radiation damage and their maximum tolerated dose (LD50) was 425-450 rad. Subcutaneous tumors grew significantly more often in young compared to adult animals (97.9% versus 69%) and this finding was not affected by TBI (96.9% versus 75%), though tumors did appear more quickly after TBI. An average of 14.7 +/- 2.8 days was required for the subcutaneous tumors to become macroscopically apparent in the adult population compared with 3.1 +/- 0.8 days in the young mice. The largest subcutaneous tumor diameter 28 days following tumor implant averaged 9.3 +/- 0.6 mm in the young animals and 5.5 +/- 1.7 mm in the adult population (P less than 0.01). Treatment of young mice with human recombinant interleukin-2 (IL-2) (10,000 Units twice a day for 28 days) produced a 27% decrease in tumor growth. This effect was abolished by prior irradiation of the young mice with 375 rad TBI. Pancreatic tumor growth also occurred more consistently in young than in adult animals (91.2% versus 64.3%) and irradiation did not affect pancreatic tumor take in either group. Occasionally intrapancreatic tumor growth was associated with liver metastases in animals that were killed after 28 days (17.8% in young and 22.2% in adult animals). However, when more than 45 days elapsed before sacrificing the animals, the incidence of hepatic metastases increased to 57.1%. This was slightly less than the incidence of hepatic lesions found after direct injection of cancer cells into the liver by portal vein injection (71.4%). Direct extension of tumor into surrounding tissues was common with frequent involvement of the duodenum (83.7%), kidneys (30.6%), and other intraabdominal organs (43.9%). Survival was significantly longer in adult compared to young mice.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of intermittent exposure and drug-free intervals on the in vitro vascular tolerance to nitroglycerin.

Recent reports suggest that intermittent nitroglycerin (NTG) treatment, incorporating a daily nitrate-free interval, can avoid the tolerance associated with continuous NTG therapy. This study has investigated whether an in vitro model of NTG tolerance could be used to examine the mechanisms of tolerance avoidance (by intermittent NTG exposure) and tolerance reversal. Isolated rat abdominal aortic rings were exposed to 55 microM NTG at varying intervals over a 60 min period, and the concentration-relaxation curves to NTG were subsequently determined. Intermittent NTG exposure (either 12 x 0.5 min or 3 x 2 min) significantly reduced NTG tolerance compared to continuous exposure over the same period of time (60 min). The diminished tolerance was apparently due to the reduced total exposure time, since the NTG responsiveness of aortic rings exposed to NTG intermittently or continuously for 6 min of the incubation period was not significantly different. Under the present conditions, in vitro NTG tolerance could be reversed if sufficient washout time was allowed. Thus, aortic rings exposed to NTG for the initial, but not the final, 6 min of incubation were not tolerant to NTG. In addition, rings exposed to NTG for 3 min exhibited near-maximal tolerance after 5 min washout, but no tolerance after 60 or 120 min washout. It appears, therefore, that the isolated vessel retains the "repair" mechanism responsible for tolerance reversal under the present conditions. This study suggests that the in vitro model of NTG tolerance may be useful for investigating the characteristics and mechanisms of tolerance avoidance and reversal, as well as tolerance induction.

Cellular mechanisms of nitrate action.

It is now generally accepted that organic nitrates generate their vasodilator action via production of nitric oxide. However, the cellular location of the metabolic enzyme(s) responsible for such conversion has not been defined. We examined the production of nitric oxide, via chemiluminescence detection, by various cellular fractions of the bovine coronary artery. We were able to show that the highest activity resides in the plasma membrane. Future isolation and characterization of such metabolic systems will greatly assist our understanding of nitrate action and tolerance. Several cellular mechanisms for nitrate tolerance have been proposed. Among the most popular theories is the "intracellular sulfhydryl depletion hypothesis" originally proposed by Needleman et al. The primary supportive data for this mechanism are that exogeneously added thiols (such as N-acetylcysteine) can potentiate the in vivo activity of nitroglycerin and can partially reverse nitrate tolerance. We showed that a cellular-impermeant thiol, viz: glutathione, can also potentiate the hemodynamic effect of nitroglycerin in rats. We subsequently showed that exogenously administered thiols can promote the formation of vasoactive S-nitrosothiols in blood. Thus, the beneficial effects of thiols on nitrate action might be mediated through an extracellular pathway. Another cellular mechanism for nitrate tolerance suggested that tolerance is caused by an alteration of the enzyme, guanylate cyclase. We showed, however, that blood vessels made tolerant to nitroglycerin remain fully responsive (in terms of in vitro relaxation) toward nitric oxide and S-nitrosothiols. These data showed that, as far as relaxation is concerned, nitrate tolerance did not cause a significant alteration of guanylate cyclase activity toward nitric oxide and S-nitrosothiols.

Mechanisms for the pharmacologic interaction of organic nitrates with thiols. Existence of an extracellular pathway for the reversal of nitrate vascular tolerance by N-acetylcysteine.

Recent reports have shown that the coadministration of N-acetylcysteine (NAC) potentiated the hemodynamic actions of i.v. nitroglycerin (NTG) and reversed NTG tolerance in humans. This study has investigated the feasibility of various pharmacokinetic and biochemical mechanisms for the thiol-organic nitrate interaction, using the rat as an animal model. In order to establish that the potentiating interaction between NAC and NTG can be reproduced in the rat, NTG dose-blood pressure response curves were determined before and during concurrent thiol infusion. The hypotensive effect of NTG was enhanced significantly by NAC and glutathione, but not by N-acetylserine, showing clearly that the potentiating effect of NAC was due specifically to its thiol functional group. The systemic clearance of NTG was not affected significantly by NAC coinfusion. In addition, the intracellular metabolism of NTG in thoracic aorta segments from rats infused previously with NAC or N-acetylserine was similar, both with respect to total production of metabolites and their distribution. Thus, the enhancement of NTG action could not be attributed apparently to an effect of NAC on NTG systemic pharmacokinetics or vascular metabolism of NTG. Because glutathione, which does not enter cells readily, also potentiated the effects of NTG, the possibility of an extracellular pathway for the thiol-organic nitrate interaction was examined. In vitro degradation of NTG in plasma and blood was accelerated in the presence of NAC (or glutathione). NAC also promoted the formation of S-nitroso-N-acetylcysteine from NTG in rat and human plasma and human blood.(ABSTRACT TRUNCATED AT 400 WORDS)

Differential stimulation of synaptosomal norepinephrine uptake by high salt diet in Dahl rats.

We have studied the norepinephrine (NE) uptake processes directly in synaptosomes isolated from the hypothalamus of both Dahl salt-sensitive (DS) and Dahl salt-resistant (DR) rats. Both DS and DR rats were divided into two dietary groups, one high salt diet group and one low salt diet group. NE uptake was highly sodium dependent (averaging 80%) and ouabain sensitive (averaging 55%). The initial 3H-NE uptake by the hypothalamic synaptosomal fraction of DR and DS rats on a low salt diet during the first 10-min incubation period averaged 1.19 +/- 0.083 and 1.50 +/- 0.138 pmol/mg protein respectively while those of DR and DS on a high salt diet were 1.69 +/- 0.124 and 1.64 +/- 0.092 pmol/mg protein respectively. Baseline values of NE uptake on low salt diet were relatively high in DS compared to that in DR controls. High salt diet had a stimulatory effect on the net uptake of 3H-NE by hypothalamic synaptosomes of both strains of rats, DS showed an overall enhancement of 9% as compared to DR (42% increase, P = 0.003). This differential enhancement by the high salt diet was apparently contributed to by the sodium-mediated and ouabain sensitive amine uptake process and possibly resulted from a defective inducibility of (Na+-K+)-ATPase in DS rats.