Subgroup analyses of a randomized trial of DHA supplementation for infants born preterm with assessments of cognitive development up to 7-years of age: What happens in infants born <29 weeks' gestation?

A recent trial showed that high-dose docosahexaenoic acid (high-DHA) supplementation of infants born <29 weeks' gestation improves intelligence quotient (IQ) at five years' corrected age. However, this finding has not been detected by other trials of DHA, which either did not measure IQ or included more mature infants. We analyzed the subgroup of 204 infants born <29 weeks' from our earlier randomized trial of high-DHA (∼1 % total fatty acids) or standard-DHA (∼ 0.3 % total fatty acids). Participants were assessed for cognition at 18 months, and IQ and behavior at seven years' corrected age. No group differences were detected for mean cognitive, IQ or behavior scores. At 18 months, 18.8 % of children in the high-DHA group had a cognitive score <85, compared with 31.1 % of children in the standard-DHA group, but at seven years there was no difference. Although an underpowered post-hoc subgroup analysis, this study provides limited support to recommendations that infants born <29 weeks' gestation require supplemental DHA.

Safety of meningococcal B vaccine (4CMenB) in adolescents in Australia.

BACKGROUND: Four-component meningococcal B (4CMenB) vaccine is licensed in many countries but has had limited use in adolescents despite this age group being at increased risk of meningococcal disease. OBJECTIVES: To assess the safety profile of two doses of 4CMenB in adolescents. METHODS: Cluster randomised controlled trial of senior school students in South Australia (SA) with participating schools randomised to intervention (4CMenB) or control. Vaccine safety was monitored using the South Australian Vaccine Safety Surveillance System (SAVSS), a spontaneous reporting system for adverse events following immunisation (AEFI) with enhanced follow-up of AEFI. RESULTS: 58,637 doses of 4CMenB vaccine were administered to 30,522 students (median age 16 years) during 2017-2018. Of 18,348 and 12,174 students vaccinated in 2017 and 2018, 97.3% and 84.3%, respectively, received both scheduled doses (N = 28,115). 193 AEFI in 187 students were reported with a reporting rate of 0.32% (95%CI: 0.28-0.39%). Seventy individuals sought medical review, including nine serious adverse events. 98% (166/169) of those who were contactable for AEFI follow-up (87.6% 169/193) reported resolution of the event. Most common AEFI were injection site reaction (126/193), headache (99/193) and nausea (61/193). AEFI were more frequently reported in females (aOR = 1.409 (95%CI: 1.002, 1.980)), schools with high level of educational advantage (adjusted Odds Ratio (aOR) = 1.515 (95%CI: 1.005, 2.284)), following first dose (aOR = 1.619 (95%CI: 1.168, 2.244)), and in 2017 (aOR = 1.437 (95%CI: 1.001, 2.064)). Reported AEFI declined with increasing age (aOR = 0.771 (95%CI: 0.673, 0.883)). CONCLUSION: In this largest post-licensure use of 4CMenB in adolescents, the low AEFI reporting rate provides real-world evidence of 4CMenB safety in this age group. (ClinicalTrials.gov number: NCT03089086).

Omega-3 fatty acid supplementation in pregnancy-baseline omega-3 status and early preterm birth: exploratory analysis of a randomised controlled trial.

OBJECTIVE: To identify a polyunsaturated fatty acid (PUFA) biomarker able to detect which women with singleton pregnancies are most likely to benefit from omega-3 supplementation to reduce their risk of early preterm birth. DESIGN: Exploratory analysis of a randomised controlled trial. SETTING: Six Australian hospitals. POPULATION: Women with a singleton pregnancy enrolled in the ORIP trial. METHODS: Using maternal capillary whole blood collected ~14 weeks' gestation, the fatty acids in total blood lipids were quantified using gas chromatography. Interaction tests examined whether baseline PUFA status modified the effect of omega-3 supplementation on birth outcomes. MAIN OUTCOME MEASURE: Early preterm birth (<34 weeks' gestation). RESULTS: A low total omega-3 PUFA status in early pregnancy was associated with a higher risk of early preterm birth. Among women with a total omega-3 status ≤4.1% of total fatty acids, omega-3 supplementation substantially reduced the risk of early preterm birth compared with control (0.73 versus 3.16%; relative risk = 0.23, 95% confidence interval [CI] 0.07-0.79). Conversely, women with higher total omega-3 status in early pregnancy were at lower risk of early preterm birth. Supplementing women with a baseline status above 4.9% increased early preterm birth (2.20 versus 0.97%; relative risk = 2.27, 95% CI 1.13-4.58). CONCLUSIONS: Women with singleton pregnancies and low total omega-3 PUFA status early in pregnancy have an increased risk of early preterm birth and are most likely to benefit from omega-3 supplementation to reduce this risk. Women with higher total omega-3 status are at lower risk and additional omega-3 supplementation may increase their risk. TWEETABLE ABSTRACT: Low total omega-3 fat status helps identify which women benefit from extra omega-3 to reduce early prematurity.

Prenatal omega-3 LCPUFA and symptoms of allergic disease and sensitization throughout early childhood - a longitudinal analysis of long-term follow-up of a randomized controlled trial.

BACKGROUND: Randomized controlled trials of prenatal omega (ω-3) long chain polyunsaturated fatty acid (LCPUFA) supplementation are suggestive of some protective effects on allergic sensitization and symptoms of allergic disease in childhood. Due to the nature of the atopic march, investigation of any effects of this prenatal intervention may be most informative when consistently assessed longitudinally during childhood. METHODS: Follow-up of children (n = 706) with familial risk of allergy from the Docosahexaenoic Acid to Optimize Mother Infant Outcome (DOMInO) trial. The intervention group received fish oil capsules (900 mg of ω-3 LCPUFA) daily from <21 weeks' gestation until birth; the control group received vegetable oil capsules without ω-3 LCPUFA. This new longitudinal analysis reports previously unpublished data collected at 1 and 3 years of age. The allergic disease symptom data at 1, 3 and 6 years of age were consistently reported by parents using the "International Study of Asthma and Allergies in Childhood" (ISAAC) questionnaire. Sensitization was determined by skin prick test to age specific, common allergen extracts. RESULTS: Changes over time in symptoms of allergic disease with sensitization (IgE-mediated) and sensitization did not differ between the groups; interaction p = 0.49, p = 0.10, respectively. Averaged across the 1, 3 and 6-year assessments, there were no significant effects of prenatal ω-3 LCPUFA supplementation on IgE-mediated allergic disease symptoms (adjusted relative risk 0.88 (95% CI 0.69, 1.12), p = 0.29) or sensitization (adjusted relative risk 0.97 (95% CI 0.82, 1.15), p = 0.76). Sensitization patterns to common allergens were consistent with the atopic march, with egg sensitization at 1 year strongly associated with house dust mite sensitization at 6 years, (p < 0.0001). DISCUSSION: Although there is some evidence to suggest that maternal supplementation with 900mg ω-3 LCPUFA has a protective effect on early symptoms of allergic disease and sensitization in the offspring, we did not observe any differences in the progression of disease over time in this longitudinal analysis. Further investigation into the dose and timing of ω-3 LCPUFA supplementation, including long-term follow up of children using consistent outcome reporting, is essential to determine whether this intervention may be of benefit as a primary prevention strategy for allergic disease. CONCLUSION: Maternal supplementation with 900 mg of ω-3 LCPUFA did not change the progression of IgE-mediated allergic disease symptoms or sensitization throughout childhood from 1 to 6 years. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry (ACTRN); DOMInO trial ACTRN12605000569606, early childhood allergy follow up ACTRN12610000735055 and 6-year allergy follow up ACTRN12615000498594.

A dose response randomised controlled trial of docosahexaenoic acid (DHA) in preterm infants.

Thirty one infants born less than 30 weeks׳ gestational age were randomised to receive either 40 (n=11), 80 (n=9) or 120 (n=11) mg/kg/day of docosahexaenoic acid (DHA) respectively as an emulsion, via the feeding tube, commenced within 4 days of the first enteral feed. Twenty three infants were enroled in non-randomised reference groups; n=11 who had no supplementary DHA and n=12 who had maternal DHA supplementation. All levels of DHA in the emulsion were well tolerated with no effect on number of days of interrupted feeds or days to full enteral feeds. DHA levels in diets were directly related to blood DHA levels but were unrelated to arachidonic acid (AA) levels. All randomised groups and the maternal supplementation reference group prevented the drop in DHA levels at study end that was evident in infants not receiving supplementation. Australian New Zealand Clinical Trials Registry: ACTRN12610000382077.

U-shaped relationship between tissue docosahexaenoic acid and atrial fibrillation following cardiac surgery.

BACKGROUND/OBJECTIVES: Randomised controlled trials (RCTs) evaluating the effect of fish oil supplementation on postoperative atrial fibrillation (POAF) following cardiac surgery have produced mixed results. In this study, we examined relationships between levels of red blood cell (RBC) n-3 long-chain polyunsaturated fatty acids (LC-PUFAs) and the incidence of POAF. SUBJECTS/METHODS: We used combined data (n=355) from RCTs conducted in Australia and Iceland. The primary end point was defined as POAF lasting >10 min in the first 6 days following surgery. The odds ratios (ORs) for POAF were compared between quintiles of preoperative RBC n-3 LC-PUFA levels by multivariable logistic regression. RESULTS: Subjects with RBC docosahexaenoic acid (DHA) in the fourth quintile, comprising a RBC DHA range of 7.0-7.9%, had the lowest incidence of POAF. Subjects in the lowest and highest quintiles had significantly higher risk of developing POAF compared with those in the fourth quintile (OR=2.36: 95% CI; 1.07-5.24 and OR=2.45: 95% CI; 1.16-5.17, respectively). There was no association between RBC eicosapentaenoic acid levels and POAF incidence. CONCLUSIONS: The results suggest a 'U-shaped' relationship between RBC DHA levels and POAF incidence. The possibility of increased risk of POAF at high levels of DHA suggests an upper limit for n-3 LC-PUFAs in certain conditions.

Prevalence and determinants of age-related macular degeneration in central Sri Lanka: the Kandy Eye Study.

AIMS: To determine the prevalence, associations and risk factors for age-related macular degeneration (ARMD) in central Sri Lanka. METHODS: The study was a population-based, cross-sectional survey of residents aged > or = 40 years in rural Sri Lanka. ARMD was assessed on dilated fundoscopy using the International Age-Related Maculopathy Epidemiology Study Group classification system. RESULTS: Of the 1721 subjects identified, 1375 participated (79.9%). Of the participants, 1013 were aged > or = 50 years (73.6%). The prevalence of any ARMD (adjusted for study design) was 4.72 (95% CI 2.22 to 7.20)% with 3.82 (95% CI 1.60 to 6.04)% early ARMD and 1.70 (95% CI 0.14 to 3.27)% late ARMD. Age (p<0.001) and Sinhalese ethnicity (p = 0.016) were significantly associated with ARMD. Men had a tendency toward a higher prevalence of ARMD than women, although this was not statistically significant (p = 0.081). Ocular risk factors such as cortical cataract (p = 0.024) and pseudophakia (p = 0.003) were associated with ARMD on the univariate but not multivariate analyses. Illiteracy and the identification of social supports were significantly associated with ARMD on univariate analyses. However, only social support was statistically significant after multivariate analysis (p = 0.024). CONCLUSIONS: Although the prevalence of ARMD is slightly lower in Sri Lanka than surrounding regions, it contributes to a higher proportion of visual impairment, including blindness. Risk factors include age and Sinhalese ethnicity.

Studies of granulosa cell maturation in dominant and subordinate bovine follicles: novel extracellular matrix focimatrix is co-ordinately regulated with cholesterol side-chain cleavage CYP11A1.

During growth of antral ovarian follicles granulosa cells first become associated with a novel type of extracellular matrix, focimatrix, and at larger sizes follicles become either subordinate or dominant. To examine this, bovine subordinate (9.0+/-S.E.M. 0.4 mm; n=16), partially dominant (12.0+/-0.6 mm; n=18) and fully dominant (15.0+/-0.4 mm; n=14) follicles were examined by real time RT-PCR analyses of granulosa cells and by immunohistochemistry of focimatrix. Changes in the expression of FSH receptor, LH receptor, cholesterol side-chain cleavage (CYP11A1), 3beta-hydroxysteroid dehydrogenase, aromatase (CYP19A1) and inhibin-alpha and beta-B were observed as expected for follicle sizes examined. After adjusting for size differences, only CYP11A1 was significantly different between the groups, and elevated in dominant follicles. Also after adjusting for differences in size there were no significant differences in expression of focimatrix components collagen type IV alpha-1 (COL4A1), laminin beta-2, nidogen 1 (NID1), and perlecan (HSPG2) or the volume density of NID1 and -2 and HSPG2. The volume density of focimatrix components in laminin 111 was significantly elevated in dominant follicles. Adjusting for analysis of more than one follicle per animal and for multiple correlations, CYP11A1 mRNA levels were highly correlated with the focimatrix genes COL4A1, NID1 and -2 and HSPG2. Thus, focimatrix may potentially regulate CYP11A1 expression, and the regulation of both could be important in follicular dominance.

A partial nucleated differential cell count of the bone marrow aspirate that is independent of peripheral blood dilution.

In the bone marrow (BM) nucleated differential cell count (NDC), myeloblasts are enumerated as a percentage of total nucleated cells, which are inevitably diluted with peripheral blood nucleated cells (PBNC) during BM aspiration. We propose a partial NDC (PNDC) comprising only immature haemopoietic cells capable of division, i.e. myeloblasts, promyelocytes, myelocytes and erythroblasts. We show that the myeloid : erythroid (M : E) ratio of the PNDC remains approximately constant in progressively dilute aliquots of BM aspirates. We determined the PNDC in 22 healthy subjects and investigated the effect of peripheral blood dilution on disease stratification of 66 BM aspirates with myelodysplastic syndromes (MDS). NDC and PNDC myeloblast counts were compared and the equivalent PNDC myeloblast counts for NDC myeloblast threshold counts of 5, 10 and 20% were derived. Reclassification of MDS samples with the PNDC resulted in a change in disease category in 33.3% of 51 MDS samples with NDC myeloblast counts ranging from 3 to 26%. The PNDC is independent of PBNC dilution and can be determined in dilute BM samples. It alters the disease category in a significant proportion of BM aspirates with MDS and has the potential to better stratify MDS to improve clinical outcomes and treatment.

Prevalence of exfoliation syndrome in central Sri Lanka: the Kandy Eye Study.

AIMS: To report the prevalence and correlates of exfoliation syndrome (XFS) in central, rural Sri Lanka. METHODS: A population-based, cross-sectional ophthalmic survey of inhabitants 40 years of age and over from villages in the Kandy District was conducted. Selection was randomised using a cluster sampling process. 1721 eligible participants were identified, 1375 participated. A detailed ophthalmic history and examination including ocular biometry was made of each participant. RESULTS: The prevalence of XFS was estimated to be 1.1% (95% CI 0.5 to 1.5%; 22 eyes). XFS was bilateral in eight subjects, unilateral in six subjects. Univariate analysis demonstrated a significant association between XFS and increasing age (p<0.001), increasing intraocular pressure (odds ratio 1.2; 95% CI 1.09 to 1.27; p<0.001), nuclear cataracts (odds ratio 1.92; 95% CI 1.47 to 2.51; p<0.001), visual impairment (odds ratio 9.72; 95% CI 3.01 to 31.44; p<0.001) and a history of hypertension (odds ratio 3.89; 95% CI 1.14 to 13.16; p = 0.030). CONCLUSION: XFS in this Sri Lankan population was associated with advanced age, raised intraocular pressure, nuclear cataracts, hypertension and visual impairment.

Estrogen inhibits the vascular injury response in estrogen receptor alpha-deficient mice.

The atheroprotective effects of estrogen in women are well recognized, but the underlying mechanisms responsible are not well understood. Blood vessel cells express the classic estrogen receptor, ER alpha (ref. 2-6), and are directly affected by estrogen, which inhibits the development of atherosclerotic and injury-induced vascular lesions. We have generated mice in which the ER alpha gene is disrupted and have used a mouse model of carotid arterial injury to compare the effects of estrogen on wild-type and estrogen receptor-deficient mice. Increases in vascular medial area and smooth muscle cell proliferation were quantified following vascular injury in ovariectomized mice treated with vehicle or with physiologic levels of 17 beta-estradiol. Surprisingly, in both wild-type and estrogen receptor-deficient mice, 17 beta-estradiol markedly inhibited to the same degree all measures of vascular injury. These data demonstrate that estrogen inhibits vascular by a novel mechanism that is independent of the classic estrogen receptor, ER alpha.

Clinical results of common strategies used to revise infrainguinal vein grafts.

PURPOSE: Patients who have failing infrainguinal bypass grafts or failed grafts reopened with lytic therapy represent a group at high risk of subsequent failure. Previous studies suggest that vein patch angioplasty and jump grafting may be less durable than interposition grafting as a method of correcting graft lesions. Our objective was to assess the value of various technical strategies for graft revision in a series of autogenous infrainguinal bypass grafts and to assess how these variables might affect cumulative graft patency (CGP) rates. METHODS: We retrospectively reviewed the clinical course, anatomic sites of revision, and type of revision performed on 67 grafts in 58 patients who underwent at least one revision from 1991 to 1995. Results were assessed with regression analysis and Kaplan-Meier estimates of CGP rates (p < 0.05 was considered significant). RESULTS: Sixty-seven vein grafts underwent revision of 112 anatomical sites in 95 operations. Forty-nine of 67 grafts were single-segment greater saphenous vein grafts and 18 were composite (> 1 segment) grafts, with an overall 5-year CGP rate of 72%. No difference was observed between the 4-year CGP rate in grafts with hemodynamically significant distal anastomotic stenoses repaired primarily with jump grafts (n = 20, 71% CGP rate) and those with stenoses found only in the graft body (n = 41, 89% CGP rate). Vein patch angioplasty was used primarily, but not exclusively, for focal graft body stenoses (n = 35), whereas interposition grafts (n = 11) were reserved for more diffuse strictures; no significant difference in 3-year CGP rates was observed (94% and 73%, respectively). CONCLUSION: Using an appropriate revision strategy that favors vein patch angioplasty for graft body lesions and jump grafts for distal anastomotic lesions, acceptable assisted patency rates can be achieved in grafts that are at risk for repeated failure.

The challenge of HIV prevention among high-risk adolescents.

This article reports findings from an exploratory study of HIV knowledge and risk behaviors among 60 teenagers and young men engaged in the street life of Hollywood, California. The sample was composed largely of youths of homosexual or bisexual orientation who were substance abusers, prostitutes, or both. The data suggest that although community-based education efforts may be associated with lower-risk behavior among this population, the overall risk profiles of these socially marginalized youths remained high. Inferences are drawn about the cofactors of risk that must be addressed and the education needed to enhance the health prospects of these youths.

PIP: This article opens with a review of the literature on research on HIV risk among adolescents in the US, which is organized into the following topics: adolescents and social contexts, social marginalization, sexual orientation, and developmental factors. The article then presents results of an exploratory study of HIV knowledge and risk behaviors found in 60 teenagers and young men 13-29 years old who were part of the street culture in Hollywood, California. Potential participants were included in the sample if they exhibited any combination of the following risk factors: unstable housing, involvement with illicit drugs, engagement in prostitution, or identification as gay or bisexual. Data were collected through 20-minute long structured interviews. It was found that 60% were White, 30% Black, 5% Hispanic, and 5% other; 90% were gay or bisexual; 80% were not traditionally employed; 25% were registered in school; 44% had no stable housing arrangement; and 57% used drugs. While 76% could identify a correct source of HIV infection, 75% of these also mentioned an incorrect source. Means of prevention mentioned were condom use (82%), not sharing needles (42%), and reducing the number of sexual partners (13%), but this knowledge did not translate into safe behavior (19% shared needles, 53% had intercourse without a condom, and 67% had multiple sex partners). Consistent condom use was reported by only 20%, 7% were celibate, and 63% had not undergone HIV testing. It was concluded that youth living at this level of marginalization are at very high risk of contracting HIV/AIDS and are unlikely to be served by available health promotion strategies. The distinct developmental needs of this population must be recognized in order to design programs to protect them from HIV/AIDS.

Estrogen inhibits the response-to-injury in a mouse carotid artery model.

The atheroprotective effects of estrogen are well documented, but the mechanisms responsible for these effects are not well understood. To study the role of physiologic (nanomolar) estrogen levels on the arterial response-to-injury, we applied a mouse carotid artery injury model to ovariectomized C57BL/6J mice. Mice were treated with vehicle (-E2, n = 10) or 17 beta-estradiol (+E2, n = 10) for 7 d, subjected to unilateral carotid injury, and 14 d later contralateral (normal = NL) and injured carotids from -E2 and +E2 animals were pressure fixed, harvested, and analyzed by quantitative morphometry. E2 levels in +E2 mice were consistently in the nanomolar range (2.1-2.5 nM) at days 0, 7, and 14. At 14 d, measures of both intimal and medial area were markedly increased in the -E2 group: (-E2 vs NL, P < 0.05 for both), but were unchanged from normal levels in the +E2 group (+E2 vs NL, P = NS and +E2 vs -E2, P < 0.05 for both). Cellular proliferation, as assessed by bromodeoxyuridine (BrdU) labeling, was significantly increased over NL in the -E2 mice, but this increase was markedly attenuated in the estrogen replacement group (total BrdU positive cells/section: NL = 6.4 +/- 4.5; -E2 = 113 +/- 26, +E2 = 40 +/- 3.7; -E2 vs NL, P < 0.05; +E2 vs NL, P = NS; -E2 vs +E2, P < 0.05). These data (a) demonstrate significant suppression of the mouse carotid response-to-injury by physiologic levels of estrogen replacement; (b) support the utility of this model in the study of the biologic effects of estrogen on the vascular-injury response; and (c) suggest a direct effect of estrogen on vascular smooth muscle cell proliferation in injured vessels.

Obstacles to effective child welfare service with gay and lesbian youths.

Agencies attempting to develop effective child welfare services for gay and lesbian youths must strive for effectiveness within a policy context that is politically polarized and generates more obstacles than directions. This article argues for a reconceptualization of service delivery that begins with a recognition of the unique developmental challenges facing sexual minority youths and proceeds to an examination of the systemic obstacles to providing competent services in their behalf. An ecological perspective informs the connections between developmental considerations, service issues, and human rights questions.

Effect of local acid-base status on gastric mucosal blood flow and surface cell injury by bile acid.

Topical application of 5 mM sodium taurocholate (5 TC, pH 1.2) to canine gastric mucosa results in luminal hydrogen ion (H+) loss and surface epithelial cell (SEC) injury. However, gross mucosal injury does not occur because of a protective increase in gastric mucosal blood flow (GMBF). The mechanism of this blood flow response is unknown. To test the hypothesis that mucosal acid-base status influences mucosal blood flow and surface cell injury, three groups of dogs with vascularized chambered gastric mucosae were studied during two sequential 30-min periods. Mucosae were exposed during period I to topical acidified isotonic saline (ATS, pH 1.2), and during period II to topical 5 TC. During both periods, group A (n = 5) received close intraarterial (ia) NaCl (0.15 M), group B (n = 5) close ia HCO3 (0.32 M), and group C (n = 4) close ia HCl (0.2 N). Parameters evaluated during both ATS and 5 TC periods included the luminal accumulation of DNA (DNAE, a sensitive marker of SEC exfoliation), luminal H+ loss, and GMBF measured using radiolabeled microspheres. Gastric venous pH was also measured. It was found that, compared to the NaCl group, the HCO3 group had no increase in GMBF after 5 TC exposure. Simultaneously, however, SEC loss was reduced by 48%, 604 +/- 72 with NaCl versus 314 +/- 59 micrograms/30 min DNA with HCO3, P < 0.025. Infusion of ia HCl generated a large increase in GMBF without an increase in SEC injury compared to ia NaCl. Thus, mucosal acid-base status is an important modulator of mucosal blood flow which is itself critically important to gastric mucosal protection during bile acid induced injury.

Selective lipoxygenase inhibitor reduces bile acid-induced gastric mucosal injury.

Leukotriene receptor blockade attenuates topical bile acid-induced gastric mucosal injury, suggesting that peptidyl-leukotrienes may be mediators of this injury. The purpose of this study was to test the hypothesis that a selective 5-lipoxygenase inhibitor protects against bile acid-induced gastric epithelial injury in the rat. Prior to injury with 10 and 20 mM acidified taurocholate (pH 1.2), rat stomachs were pretreated with either vehicle or WY50295K (selective 5-lipoxygenase inhibitor, 20 mg/kg). Injury was assessed by measuring net transmucosal hydrogen ion flux, luminal appearance of DNA, and gross mucosal injury. Topical 5-lipoxygenase inhibitor significantly reduced luminal H+ ion loss, surface epithelial cell loss (as measured by luminal accumulation of DNA), and gross mucosal injury in bile acid-injured stomachs compared to controls. This study lends further support to the hypothesis that leukotrienes may be mediators of bile acid-induced gastric mucosal injury.

Effect of capsaicin on gastric mucosal injury and blood flow following bile acid exposure.

Topical bile acid at low pH stimulates gastric mucosal blood flow (GMBF), thereby limiting injury to surface epithelial cells (SEC). Capsaicin-sensitive afferent neurons (ASN) are possible mediators of the GMBF response and, therefore, of mucosal protection. In order to investigate the effect of topical capsaicin (ASN stimulant) and topical lidocaine (ASN inhibitor) on SEC exfoliation and GMBF, vascularized wedges of canine gastric corpus were mounted in lucite chambers. Mucosae were pretreated for 15 min with saline (NSS), 160 microM capsaicin (CAP), 4% lidocaine (LIDO), or CAP and LIDO, followed by a 30-min exposure to acid test solution (ATS; pH 1.2). The same mucosae were then pretreated in an identical fashion followed by a second 30-min exposure to 5 mM taurocholate (5 TC; pH 1.2). Parameters evaluated during both ATS and 5 TC periods were the luminal accumulation of DNA (DNAE, a sensitive marker of SEC exfoliation) and GMBF measured using radiolabeled microspheres. It was found that, relative to NSS pretreatment, CAP pretreatment increased GMBF and decreased DNAE during exposure to both ATS and 5 TC. LIDO blocked the CAP effect on GMBF but not on DNAE. Thus, ASN stimulation by CAP enhances GMBF and is protective. ANS inhibition blocks CAP's GMBF increase but not its protective capabilities. Therefore, augmentation of GMBF is not the only mechanism by which ASNs blunt SEC exfoliation.

Nile red staining of lysosomal phospholipid inclusions.

We have employed the fluorescent dye nile red to distinguish between normal cells and cells containing lysosomal accumulations of phospholipids. When fibroblasts from an individual with a genetic deficiency in lysosomal sphingomyelinase activity (Niemann-Pick disease) were stained with nile red and visualized by fluorescence microscopy, orange-colored inclusions were observed throughout the cytoplasm. The orange fluorescent bodies could be distinguished from the neutral lipid droplets that fluoresce a brilliant yellow-gold in the presence of nile red. These inclusions were also observed in alveolar macrophages obtained from rats treated with amiodarone, an antiarrhythmic agent known to produce lysosomal phospholipidosis. Flow cytofluorometric analysis revealed that staining of these phospholipid-rich macrophages with nile red can distinguish them from control alveolar macrophages. These results demonstrate that nile red can be employed for the rapid staining of cellular phospholipid inclusions.

Hepatic lipase in the rat ovary. Ovaries cannot synthesize hepatic lipase but accumulate it from the circulation.

Hepatic lipase is proposed to have a role in steroidogenesis through its involvement in the metabolism of high density lipoproteins. We examined the activity, synthesis, distribution, and uptake of this enzyme and assessed the content of its mRNA in luteinized ovaries. We found that during peak steroidogenesis, ovaries of pregnant mare's serum gonadotropin-human chorionic gonadotropin-treated immature rats contained heparin-releasable hepatic lipase-like activity which was neutralized in a dose-dependent manner by purified antibodies to hepatic lipase isolated from post-heparin perfusates of rat livers. Quantitative immunoelectron microscopy revealed that ovarian hepatic lipase occurred along endothelial cells and was 3-fold more abundant in blood vessels of corpora lutea than those of stroma. However, hepatic lipase was not synthesized by the ovary since radiolabeled enzyme was not immunoisolated from the medium of dispersed luteinized granulosa cells incubated with [35S]methionine whereas it was present in the medium of control cells (hepatocytes). Similarly, hepatic lipase mRNA was detectable in liver but not ovaries or kidneys by Northern or slot blot analyses or by the polymerase chain reaction. Finally, 125I-labeled hepatic lipase injected into tail veins was quickly cleared from the systemic circulation, accumulating in liver, ovaries, kidneys, and spleen. Subsequent heparin injection caused rapid reappearance of radioactivity in the bloodstream and a marked decline of radiolabel in liver and ovaries but a modest decrease of that in kidneys and none in spleen. Exogenous 125I-bovine serum albumin also accumulated in all four organs but was not displaced from liver or ovaries by subsequent administration of heparin. Taken together, these data suggest that steroidogenically active ovaries possess but do not synthesize hepatic lipase. Instead, hepatic lipase originating elsewhere, presumably in the liver, is accumulated from the circulation at heparin-sensitive sites in ovarian blood vessels.