Resuscitation skills after Helping Babies Breathe training: a comparison of varying practice frequency and impact on retention of skills in different types of providers.

Background: Helping Babies Breathe (HBB), a basic neonatal resuscitation curriculum, improves early neonatal mortality in low-resource settings. Our goal was to determine retention of resuscitation skills by different cadres of providers using the approved HBB Spanish translation in a rural clinic and community hospital in Honduras. Methods: Twelve clinic and 37 hospital providers were trained in 1 d HBB workshops and followed from July 2012 to February 2014. Resuscitation skills were evaluated by objective structured clinical evaluations (OSCEs) at regular intervals. Clinic providers practiced monthly, whereas hospital providers were randomized to monthly practice for 6 months vs three consecutive practices at 3, 5 and 6 months. Results: In the rural clinic, follow-up OSCE assessment showed rapid loss of skills by 1 month after HBB training. For all providers, repeated monthly testing resulted in improvements and maintenance of OSCE performance. In the community hospital, over all time points, the group with monthly OSCEs had 2.9 greater odds of passing compared with the group who practiced less frequently. Physicians were found to have 4.3 times greater odds of passing compared with nurses. Conclusions: Rapid loss of resuscitation skills occurs after an initial training. Repeated practice leads to retention of skills in all types of providers. Further investigation is warranted to determine the clinical correlation of neonatal outcomes after HBB training.

Improvements in the delivery of resuscitation and newborn care after Helping Babies Breathe training.

BACKGROUND: To evaluate changes in neonatal resuscitation and postnatal care following Helping Babies Breathe (HBB) training at a community hospital in rural Honduras. We hypothesized that HBB training would improve resuscitation and essential newborn care interventions. METHODS: Direct observation and video recording of delivery room care spanned before and after an initial HBB workshop held in August 2013. Rates of essential newborn care interventions were compared in resuscitations performed by individuals who had and had not received HBB training, and run charts recording performance of newborn care practices over time were developed. RESULTS: Ten percent of deliveries (N=250) were observed over the study period, with 156 newborn resuscitations performed by individuals without HBB training, compared to 94 resuscitations performed by HBB trainees. After HBB training, significant improvements were seen in skin-to-skin care, breastfeeding within 60 min of age, and delayed cord clamping after 1 min (all P<0.01). More babies cared for by HBB trainees received basic neonatal resuscitation such as drying and stimulation. Run charts tracking these practices over time showed significant improvements after HBB training that were sustained during the study period, but remained below ideal goals. With improvement in drying/stimulation practices, fewer babies required bag/mask ventilation. CONCLUSION: In a rural Honduran community hospital, improvements in basic neonatal resuscitation and postnatal essential newborn care practices can be seen after HBB training. Further improvements in newborn care practices may require focused quality improvement initiatives for hospitals to sustain high quality care.

Lobe-specific increases in malondialdehyde DNA adduct formation in the livers of mice following infection with Helicobacter hepaticus.

Helicobacter hepaticus infection is associated with chronic hepatitis and the development of liver tumours in mice. The underlying mechanism of this liver carcinogenesis is not clear but the oxidative stress associated with H. hepaticus infection may result in induction of lipid peroxidation and the generation of malondialdehyde. Malondialdehyde can react with deoxyguanosine in DNA resulting in the formation of the cyclic pyrimidopurinone N-1,N(2) malondialdehyde-deoxyguanosine (M1dG) adduct. This adduct has the potential to cause mutations that may ultimately lead to liver carcinogenesis. The objective of this study was to determine the control and infection-related levels of M1dG in the liver DNA of mice over time, using an immunoslot-blot procedure. The level of M1dG in control A/J mouse livers at 3, 6, 9 and 12 months averaged 37.5, 36.6, 24.8 and 30.1 adducts per 10(8) nucleotides, respectively. Higher levels of M1dG were detected in the liver DNA of H. hepaticus infected A/JCr mice, with levels averaging 40.7, 47.0, 42.5 and 52.5 adducts per 10(8) nucleotides at 3, 6, 9 and 12 months, respectively. There was a significant age dependent increase in the level of M1dG in the caudate and median lobes of the A/JCr mice relative to control mice. A lobe specific distribution of the M1dG adduct in both infected and control mice was noted, with the left lobe showing the lowest level of the adduct compared with the right and median lobes at all time points. In a separate series of mice experimentally infected with H. hepaticus, levels of 8-hydroxy-deoxyguanosine were significantly greater in the median compared with the left lobe at 12 weeks after treatment. In conclusion, these results suggest that M1dG occurs as a result of oxidative stress associated with H. hepaticus infection of mice, and may contribute to liver carcinogenesis in this model.

Age-related alterations in 32P-postlabeled DNA adducts in livers of mice infected with the tumorigenic bacterial pathogen, Helicobacter hepaticus.

Helicobacter hepaticus causes chronic active hepatitis and liver tumors in mice, with associated increase in reactive oxygen species. Indigenous (I)-compounds are bulky DNA adducts present at low levels and detected by 32P-post-labeling. Some may be caused by reactive oxygen species; others occur normally and decrease during liver tumorigenesis. The identity of most is unknown. We investigated whether mouse liver infection by H. hepaticus and resulting progression of hepatic lesions would be associated with qualitative or quantitative changes in I-compounds. Mice were 3, 6, 9, and 12 months of age; liver disease ranged from minimal through marked. In control A/J mice, up to 20 I-compounds were detected, and the total level of these did not change with age, whereas 11 individual I-compounds showed marked age-related differences. These appeared to be coordinately regulated, as the total of these 11 adducts was constant at 6-12 months. In A/JNCr mice naturally infected with H. hepaticus, up to 12 hepatic I-compounds were found. Total levels varied markedly with age and were high at 6 and 12 months. Neither total adduct levels, nor the amount of any individual adduct, correlated positively with severity of hepatic lesions; in some cases, highest levels were found in livers with least disease. Thus, liver infection and tumorigenesis by H. hepaticus was not associated with an increase in any 32P-postlabeled DNA adduct. Marked, and distinct, age-related changes in total or individual adducts in control and infected mice suggest a role in the physiological alterations of aging and in host response to infection.

DNA adducts of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) in fetal tissues of patas monkeys after transplacental exposure.

Transplacental genotoxicity of the heterocyclic amine food-derived mutagen/carcinogen 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) has been investigated by (32)P-postlabeling assay for IQ-DNA adducts in maternal liver, placenta, and several fetal tissues of patas monkeys, after exposure to 15, 35, or 50 mg/kg IQ near the end of gestation or to the highest dose in the first or second trimester. Dose-dependent adduct formation occurred in all tissues, with the highest levels occurring in maternal liver. Adduct amounts were similar among fetal tissues and placenta, except for lower levels in fetal brain and slightly more adducts in fetal liver. Adducts in placenta, fetal liver, lung, kidney, skin, and adrenal gland, but not in maternal liver or fetal brain, increased significantly as gestation progressed. Pretreatment with phenobarbital, which induces CYP enzymes that detoxify IQ, decreased adducts in maternal liver and possibly placenta, but not in fetal tissues. The CYP inducer beta-naphthoflavone caused a significant increase in IQ-DNA adducts in fetal lungs. Regression analysis suggested that IQ activation in maternal and fetal liver and possibly placenta contributed to adduct formation in fetal tissues; adducts in placenta and/or fetal liver were strong predictors for those in most fetal tissues. The results indicate that exposure of pregnant primates to IQ results in DNA adduct formation in most fetal tissues, especially late in gestation; that upregulation of maternal detoxification does not provide fetal protection; and that adducts in placenta indicate adduct levels in fetal tissues.

Increased urinary excretion of 2-hydroxyestrone but not 16alpha-hydroxyestrone in premenopausal women during a soya diet containing isoflavones.

Asian diets high in soy are associated with lower risk for breast cancer compared with Western diets. Moreover, higher levels of two putative carcinogenic metabolites of 17beta-estradiol, 4- and 16alpha-hydroxyestrogen, and lower amounts of anticarcinogenic metabolites, 2-hydroxyestrogens, have been associated with greater breast cancer risk. In this study, we tested the hypothesis that consumption of a soya diet containing the weakly estrogenic isoflavones genistein and daidzein may alter the metabolism of 17beta-estradiol to 2- and 16alpha-hydroxylated products. Eight pre-menopausal women were placed on a soya-containing, constant diet in a metabolic unit. The diet provided 400 kilocalories from soymilk and 113-202 mg/day (158 +/- 26 mg/day, mean +/- SD) isoflavones daily for a complete menstrual cycle. After a washout period of 4 months, the subjects consumed the same diet, but with soymilk that contained <4.5 mg/day isoflavones ("isoflavone-free"). Urine samples were collected for 24 h daily for the entire cycle during each soya diet period for the analysis of daidzein, genistein, and 2- and 16alpha-hydroxyestrone. Subjects excreted measurable amounts of daidzein (11.6-39.2 mg/day) and genistein (2.9-18.2 mg/day) during the isoflavone-rich soya diet but not during the isoflavone-free soya diet. The diet rich in isoflavones increased the cycle mean daily urinary excretion of 2-hydroxyestrone (averaged over the entire cycle) from 11.6 +/- 2.06 to 17.0 +/- 2.96 nmol/12-h (P = 0.03), a 47% increase. However, the mean daily excretion of 16alpha-hydroxyestrone did not change (7.0 +/- 1.14 nmol/12-h during the isoflavone-free and 7.7 +/- 1.25 nmol/12-h during the isoflavone-rich diet; P = 0.36). The ratio of 2-hydroxyestrone to 16alpha-hydroxyestrone was higher during the isoflavone-rich soya diet (2.6 +/- 0.34) than during the isoflavone-free diet (2.0 +/- 0.32; P = 0.01), a 27% increase. These results suggest that soya isoflavones increase the metabolism of endogenous estrogens to the protective 2-hydroxylated estrogens in women, and this may play an important role in lowering 17beta-estradiol levels and the long-term risk for breast cancer.

Dietary omega-3 fatty acids as potential inhibitors of carcinogenesis: effect on DNA adduct formation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in mice and rats.

The heterocyclic amine 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is carcinogenic in the CDF1 mouse, causing lymphomas (spleen and lymph nodes) and in the F344 rat, causing mammary tumours in the female and colon tumours in the male. Dietary fish oil, a rich source of omega-3 fatty acids, exhibits chemopreventive properties in several rodent tumour models. The potential chemopreventive properties of dietary omega-3 fatty acid ethyl ester concentrate (O3C) were tested by evaluating its effects on the formation and removal of PhIP-DNA adducts. In the first experiment, a powdered AIN-76A diet containing 4.0% (w/w) O3C inhibited PhIP-DNA adduct formation in various organs of the CDF1 mouse, but not in those of the F344 rat. In a subsequent, second experiment, groups of male CDF1 mice were maintained for 43 days on AIN-76A diets containing the following percentages (w/w) of corn oil ethyl esters and O3C: 7.0 and 0, 5.5 and 1.5, 4.0 and 3.0, and 1.0 and 6.0, respectively. All animals received 0.04% (w/w) PhIP in the diet during weeks 3 and 4. Using 32P-postlabelling assays, PhIP-DNA adducts were analysed in various organs and white blood cells (WBC) on days 1, 8 and 15 after removal of PhIP from the diet. In the liver, O3C-containing diets inhibited adduct formation at all three time points (40.3-60.0%, 53.4-75.7% and 43.3-64.3% on days 1, 8 and 15, respectively). In the spleen, inhibition was evident only on days 8 (35.4-38.8%) and 15 (38.4-56.5%). O3C diets inhibited adduct formation in the stomach, small intestine and caecum at all three time points (except in the stomach and caecum on day 15) amounting to 18.5-31.5% decreases in the stomach, 40.0-60.3% decreases in the small intestine and 24.4-31.4% decreases in the caecum. The extent of inhibition was not related to O3C concentration. In the colon and WBC, adduct levels were independent of the type of diet. In all organs, adduct levels decreased significantly over time, with day 15 levels being 6.3-31.6% of those on day 1. Rate of adduct removal was independent of the type of diet. It is concluded that dietary O3C inhibits PhIP-DNA adduct formation in a target organ (spleen) as well as in non-target organs (liver and gastrointestinal tract) of the CDF1 mouse, but that the rate of adduct removal is independent of the O3C content of the diet.

Inhibition of DNA adduct formation of PhIP in female F344 rats by dietary conjugated linoleic acid.

The dietary mutagen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a mammary carcinogen in the female Fischer (F344) rat and a colon carcinogen in the male F344 rat. To exert its carcinogenicity, it is believed that PhIP needs to form adducts with DNA, a process requiring N-hydroxylation of PhIP by cytochromes P-450 1A1 and/or 1A2 (CYP 1A1 and/or 1A2), as well as further esterification of the hydroxylamine thus formed. Dietary conjugated linoleic acid (CLA) inhibits chemical carcinogenesis in various experimental models. We have examined the effect of dietary CLA on PhIP-DNA adduct formation in female F344 rats. Four-week-old animals were maintained on AIN-76A diet without or with CLA (1%, 0.5%, and 0.1% wt/wt) for 57 days. PhIP was added to the diets (0.04% wt/wt) from Days 14-42. Animals were killed (4/group) on Days 43, 50, and 57. DNA isolated from liver, mammary epithelial cells (MEC), colon, and white blood cells (WBC) was analyzed for PhIP-DNA adducts by 32P-postlabeling assays. On Day 43, CLA inhibited adduct formation in the liver (up to 58%) in a dose-dependent manner. CLA also inhibited hepatic adduct levels (29-39%) on Day 50 (at 1.0% and 0.5% CLA) and on Day 57 (53% at 0.5% CLA). CLA significantly reduced adduct levels in the WBC on Day 50 (63-70%). Adducts in MEC and the colon were not affected by dietary CLA. On Day 57, adduct levels in MEC, liver, colon, and WBC were 0-30.3%, 8.6-41.7%, 21.5-50.7%, and 7.5-11.8%, respectively, of those on Day 43. Northern blot analysis of liver RNA showed that dietary CLA did not affect steady-state levels of CYP 1A1 or 1A2 mRNA. It is concluded that dietary CLA inhibits PhIP-DNA adduct formation in liver and WBC but that those in MEC and the colon are unaffected when a low-level dietary regimen of carcinogen and inhibitor was used. In inhibiting PhIP-DNA adduct formation, CLA does not appear to act by inhibiting CYP 1A1 or 1A2 expression.

Effects of dietary conjugated linoleic acid on DNA adduct formation of PhIP and IQ after bolus administration to female F344 rats.

Meats cooked at high temperatures contain mutagenic heterocyclic amines such as 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ). In female Fischer 344 rats, IQ is a multiorgan carcinogen, whereas PhIP induces mammary adenocarcinomas. For IQ and PhIP, N-hydroxylation, catalyzed by microsomal cytochrome P-450 1A1 and/or 1A2, and then esterification, especially O-acetylation, are the principal steps leading to DNA adduct formation. Conjugated linoleic acid (CLA) is a mixture of conjugated linoleic acid isomers found in various meat and dairy products. We have examined the effect of dietary CLA on DNA adduct formation by PhIP and IQ in female Fischer 344 rats. Four-week-old animals were maintained on AIN-76A diet without or with CLA (4% wt/wt) and treated with IQ or PhIP (50 mg/kg by gavage) after two weeks. Animals were killed (4/group) one, four, and eight days later. DNA isolated from mammary epithelial cells, liver, colon, and white blood cells was analyzed for carcinogen-DNA adducts by 32P-postlabeling assays. On Day 1, dietary CLA significantly inhibited adduct formation (82.0%) in mammary epithelial cells in IQ--but not in PhIP-treated rats. In the colon, dietary CLA significantly inhibited PhIP-DNA adduct formation (18.7%) on Day 8 but increased IQ-DNA adduct formation (30.5%) on Day 8. Dietary CLA had no effect on adduct levels in liver or white blood cells. Calf thymus DNA was incubated with N-hydroxy-PhIP or -IQ in the presence of acetyl-CoA. Enzymatic activation was catalyzed by liver or mammary cytosol. A two-week pretreatment with 2% (wt/wt) dietary CLA had no effect on O-acetyltransferase-catalyzed IQ- or PhIP-DNA adduct formation. It is concluded, under certain conditions, that dietary CLA can lower IQ- and PhIP-DNA adduct formation. Overall, however, the major mode of action of CLA is probably by a mechanism other than the inhibition of the N-hydroxylation and subsequent O-acetylation of PhIP or IQ.

DNA adducts of heterocyclic amines: formation, removal and inhibition by dietary components.

The dietary mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) are carcinogenic in rodents. In F344 rats PhIP induces mammary tumors in females and colon tumors in males, while IQ induces tumors principally in the liver, Zymbal gland and intestines. In CDF1 mice, IQ induces liver, lung and forestomach tumors. We have evaluated the dynamics of formation, removal and inhibition of PhIP- and IQ-DNA adducts in these rodents. After bolus doses (50 mg/kg, by gavage) of IQ or PhIP, both IQ- and PhIP-DNA adducts were removed rapidly from both target and nontarget organs, while after 3-4 weeks of feeding IQ or PhIP (0.01-0.04%) adduct removal was much slower. Gavaging of male F344 rats with PhIP (0.1-1000 micrograms/kg/day) for 23 days resulted in accumulation of PhIP-DNA adducts in various organs, but adducts were detectable only at 100 or 1000 micrograms/kg/day. Urinary excretion of unchanged PhIP was a constant proportion (1.6-2.1%) of the daily dose over the entire dose range and was independent of duration of exposure. When weanling female F344 rats were exposed to dietary PhIP (0.01-0.04%) for 1-4 weeks, the presence of either conjugated linoleic acid (CLA; 0.1-1.0%) or indole-3-carbinol (13C; 0.1%) in the diet inhibited PhIP-DNA adduct formation (58-99%) in various organs, including the mammary gland and the colon. Similarly, the inclusion of 0.075% 4-ipomeanol (IPO) in the diet of male CDF1 mice exposed for 3 weeks to dietary IQ (0.01%) resulted in inhibition of IQ-DNA adduct formation (30-59%) in the target organs (liver, lungs, stomach) but not in a number of other organs. It is concluded that (1) the rate of PhIP- and IQ-DNA adduct removal depends on the dose and frequency of administration, (2) urinary PhIP may be a good biomarker of recent PhIP exposure and (3) CLA, I3C and IPO are potential chemopreventive agents against PhIP- or IQ-induced tumors in rodents.