We procured 4,085 daily samples of PM2.5 on Teflon filters from the Connecticut Department of Environmental Protection for six cities in Connecticut. Filters were analyzed for chemical composition, and Positive Matrix Factorization was used to determine contributions of PM2.5 sources. Risk estimates were calculated with conditional logistic regression, matching pregnancies to the same women.Methods: Birth certificates were used to select women who had vaginal singleton live births at least twice in Connecticut during 2000–2006 , 1.01 , 1.00 , and 1.09 , respectively.Results: Odds ratios of preterm birth per interquartile range increase in whole pregnancy exposure to dust, motor vehicle emissions, oil combustion, and regional sulfur PM2.5 sources and preterm birth, and the first matched analysis, that better addresses individual-level confounding potentially inherent in all past studies. There was insufficient evidence to suggest that sources were statistically significantly associated with preterm birth. However, elevated central estimates and previously observed associations with mass concentration motivate the need for further research. Future studies would benefit from high source exposure settings and longitudinal study designs, such as that adopted in this study.Conclusion: This was the first study of PMhttp://dx.doi.org/10.1289/ehp.1307741Citation: Pereira G, Bell ML, Lee HJ, Koutrakis P, Belanger K. 2014. Sources of fine particulate matter and risk of preterm birth in Connecticut, 2000–2006: a longitudinal study. Environ Health Perspect 122:1117–1122;  The Uni studies . It is u studies . A more l design . In thatts alone . Associa2.5 in Connecticut and assess the hypothesis that exposures to the anthropogenic sources are associated with elevated risk of preterm birth.The aim of this study was to identify major sources of PMStudy design and population. This was a longitudinal study of a retrospective cohort—retrospective because records were obtained after the women delivered their children, and longitudinal by repeated pregnancies to the same women over time. We investigated women’s exposure to sources of ambient PM2.5 and preterm birth across successive pregnancies in Connecticut, 2000–2006. From a population of 271,204 singleton live births without congenital anomaly we sequentially excluded 157 (0.06%) birth records with missing gestational age; 1,846 (0.86%) records that could not be geocoded; 1,505 (0.56%) birth records to women who resided farther than 40 km from a monitoring station for PM2.5 mass concentration; 1,443 (0.54%) records with missing parity; and 72,362 (27.18%) births by cesarean delivery. The remaining reference population consisted of birth records for 193,891 neonates to 152,934 women.2.5 monitor for which PM2.5 chemical components were measured , and, finally, women who gave birth at least twice during the study period . This resulted in a final study population of 48,208 neonates born to 23,123 women.For the longitudinal analyses, this reference population was further sequentially restricted to records where at least 75% of the weekly mass concentrations were available in each trimester and whole pregnancy , women who lived within 40 km of a PMData sources and variables. Birth records were obtained from the Connecticut Department of Health  for all registered births in Connecticut from 1 January 2000 to 31 December 2006. Each observation contained variables for the residential location at birth; pregnancy-related risk factors ; and sociodemographic risk factors .Outcome assessment. Preterm birth was defined as birth before 37 weeks completed gestation. Period of gestation was obtained from the birth certificate record. This was the clinical best estimate of gestational age, based on ultrasound or last menstrual period if ultrasound was not available. More than 90% of all births in Connecticut have a dating ultrasound. First and second trimesters were defined as weeks 1–13 and 14–26, respectively. Third trimester was defined as commencing week 27 and ending at the end of week 36 or birth, whichever was earlier.Sampling and chemical analysis. PM2.5 samples, collected on Teflon filters, were obtained from the Connecticut Department of Energy & Environmental Protection  for six sites  for the period 2000–2006. The 24-hr integrated PM2.5 samples were taken every third day in Bridgeport (692 samples), Danbury (667 samples), Hartford (694 samples), New Haven (686 samples), Norwalk (683 samples), and Waterbury (663 samples). These filters were originally used by the Connecticut Department of Energy & Environmental Protection to measure PM2.5 total mass according to federal U.S. Environmental Protection Agency (EPA) sampling protocol (2.5 elements using X-ray fluorescence (XRF) for 51 elements (protocol . The filelements  and optielements . XRF waselements . To ensuelements . BrieflySource apportionment. Source apportionment was conducted separately for each monitoring site. Source apportionment was conducted using Positive Matrix Factorization (PMF) with PMF 3.0 developed by the X (n samples by m species), of observed species concentrations. The result is a representation of X as the product of a matrix, G (n samples by p sources), of source contributions for each sample and a matrix, F (p sources by m species), of species profiles for each source plus a matrix of residuals. The matrix of residuals, E (n samples by m species), contains the differences between the observed and modeled concentrations of each species for each sample.X = GF + E. [1]G and F are constrained to be non-negative and minimize the sum of the squared residuals scaled by their standard deviation (uncertainty), Q:Elements of Q = Σ∀ij [(X – GF)ij/Σij]2, [2]i = 1, 2,…, n (samples) and j = 1, 2,…, m (species).where The PMF solution was rotated, with FPEAK parameters within the interval  . The resExposure assessment. Weekly mean concentrations of the identified PM2.5 source contributions  were calculated as 7-day averages of the daily source contributions for each monitor over the 2000–2006 period. Mean exposures were computed for each week of gestation and then used to compute exposure for each trimester and whole pregnancy. Only exposure estimates to week 36 were included in third trimester and whole pregnancy. Given previously observed associations between preterm birth and PM2.5 mass concentration, we assigned exposure estimates to each woman based on the closest monitor within 40 km of the women’s residence at time of birth . Separate models were fitted for each trimester and whole pregnancy exposure. Adjustment was made for average number of cigarettes smoked per day , maternal age , and parity  for each pregnancy. Adjustment was made for these factors because of their potential to change considerably between pregnancies, and they are strong independent risk factors for preterm birth  choice of the 40-km buffer distance, b) smoking status, c) certainty of the LMP  date, d) changing residential address between pregnancies, e) changing residential address within pregnancies, f) exclusion of births by cesarean section, g) unmeasured temporal confounders, h) adjustment for additional socioeconomic variables .Sensitivity of analyses to buffer distance was investigated by repeating analyses after restriction to women living within 10 km, 20 km, and 30 km from a monitor. Smoking status during pregnancy was ascertained with a binary variable (yes/no). Certainty of LMP date was also ascertained with a binary variable (certain/uncertain). We identified women who changed address between pregnancies using address recorded at delivery, and those who changed address during pregnancy with a variable also recorded at delivery on the number of months the women has resided at the address. We calculated a propensity score for preterm birth conditional on a GAM spline of conception date with an unmatched logistic regression model using the reference population, where the smoothing parameters were selected by generalized cross-validation. This spline term was then used as an adjustment term in the longitudinal analysis to account for potential unmeasured temporal confounders. Sensitivity to additional socioeconomic adjustment was investigated with marital status at the individual level and percent unemployed and median household income at the census-tract level.Approvals. Institutional review board approvals for this study were obtained from the Yale Human Investigation Committee and the Connecticut Department of Health.Preterm birth rate and characteristics of the longitudinal study population. The preterm birth rate was 7.45% for the original source population before exclusions, 6.40% for the reference population, and 5.36% for the longitudinal study population . The longitudinal study design resulted in few women ≥ 40 years of age  and more women < 20 years of age . Fewer women had < 12 years of education . Most women were married , white , nonsmokers , and had no other children . A small proportion of women delivered only preterm neonates . There were 1,966 (8.50%) women who delivered both preterm and term neonates.2.5Identified sources of PM. The proportion of PM2.5 component observations below the MDL varied by site . The proportion of variation (R2) in observed PM2.5 explained by the PMF solution was 86% in Bridgeport, 92% in Danbury, 88% in Hartford, 74% in New Haven, 92% in Norwalk, and 92% in Waterbury. The source apportionment resulted in detection of five sources  in Bridgeport, Danbury, Hartford, and New Haven.2.5Spatial variation in sources of PM. In general, the factor profiles, which reflect the composition of emissions in each source, , whereas both motor vehicle emissions and oil combustion peaked in the cold season (December–February) (ebruary) . Briefly2.5Exposure to anthropogenic sources of PM. Throughout pregnancy, women were exposed to higher median levels of regional sulfur (4.26 μg/m3) than of motor vehicle emissions (3.52 μg/m3), dust (1.49 μg/m3), and oil combustion (1.29 μg/m3)  . For eac2.5 and preterm birthAssociation between anthropogenic sources of PM. Among all women, the odds ratios (ORs) for preterm birth for IQR increases in whole pregnancy exposure to dust, motor vehicle emissions, oil combustion, and regional sulfur were 1.01 , 1.01 , 1.00 , and 1.09 , respectively  increase in first- and third-trimester exposure to regional sulfur were 1.06  and 1.03 , respectively, with some suggestion of stronger associations among Hispanic women. For exposure to oil combustion, adverse associations were strongest for elevated exposure in first trimester, more prominently among Hispanic  and black women . Among Hispanic women, associations in the opposite direction (OR < 1) were observed for IQR increases in second-trimester exposure to oil combustion  and dust .Sensitivity analyses. The original effect estimates were generally robust to choice of the 40-km buffer distance , the inclusion of smokers in the study population , accuracy of the LMP date , change of residential address either between  or within  pregnancies, adjustment for unmeasured temporal confounders , and adjustment for additional socioeconomic variables . We excluded all cesarean sections from original analyses because final gestational length for cesarean sections without labor is determined before delivery, and Connecticut birth certificates were not suitable for separately identifying cesarean sections that occurred with labor and those that occurred without labor. It is plausible that a fraction of morbidities antecedent of cesarean section were caused by exposure to PM2.5 sources, in which case births by cesarean section would not be excluded. However, exposure would be misclassified for all of these individuals in our study because we did not have information on the time of onset/diagnosis of the antecedent during pregnancy. Nonetheless, the original results were robust to the exclusion of births by cesarean section .Summary of results. By matching pregnancies to the same woman, we investigated the hypothesis that women are more likely to deliver preterm when exposure to anthropogenic sources of PM2.5 is elevated. This was achieved by longitudinal follow-up of outcomes among > 23,000 women over a 7-year period. This approach lowers the considerable potential for confounding by sociodemographic factors, a limitation of previous studies that relied on between-women comparisons. We also procured > 4,000 daily samples of PM2.5 from the Connecticut Department of Energy & Environmental Protection for six cities in Connecticut, which significantly improved both the temporal resolution as well as the geographic relevance of the identified sources of exposure.The results demonstrated that the greatest contributions to exposure were attributed to regional sulfur and motor vehicle emissions. Concentrations of these sources were more than double the concentrations of sea salt, oil combustion, and dust. The results also showed temporal variation in sources, with peak regional sulfur in the warm season (June–August). Motor vehicle emissions and oil combustion peaked in the cold season (December–February). There was insufficient evidence to conclude that fine particulate levels of dust, motor vehicle emissions, oil combustion, and regional sulfur were associated with risk of preterm birth. Although central estimates for three of the four sources were positive (and the fourth null), they were not statistically significant. Adjusted ORs per IQR increase in whole pregnancy exposure to dust, motor vehicle emissions, oil combustion, and regional sulfur were 1.01 , 1.01 , 1.00 , and 1.09  respectively.p = 0.130) relate to the adverse associations observed in past studies  per IQR increase in diesel PM2.5 in Los Angeles, California  we investigated individual-level risk rather than its approximation from between-women comparisons more susceptible to residual socioeconomic confounding; b) the California study investigated a population living within 8 km of a monitoring site, who tend to have worse risk profiles than the general population ; or c) whole pregnancy exposure to PM2.5 (mass concentration) was higher in the California study  than this study .lifornia . That stthan CMB . Our resPotential causal pathways and causal inference. Pathways by which air pollution might cause preterm birth might include preeclampsia  because the longitudinal nature of the study limited sample size. Based on the interquartile range of the interbirth interval distribution of women who delivered a child in the first year of the study period, we would expect to have observed a subsequent birth (if it occurred) for half to three-quarters of women using a study period of 7 years. Exclusion of women with longer interbirth intervals might have induced a degree selection bias, for example, if PM2.5 sources contributed to subfertility, and subsequently longer interbirth intervals and fertility treatment–associated preterm birth. Exposures did not differ between the study, reference, and source populations . A further limitation of this study was that exposure was assigned based on residential address recorded at delivery. Exposure would be misclassified by residential movement during pregnancy and exposures away from the home. However, sensitivity analyses provided greater confidence that estimates were robust to residential movement . Finally, future longitudinal studies would benefit from additional information on risk factors for preterm birth that tend to change between pregnancy , although we expect most to be nonassociated with exposure.2.5 sources on preterm birth and the first study to make use of longitudinal comparisons (matching on the mother) to better address individual-level confounding potentially inherent in all past studies. The greatest contributions to total mass of fine particulate matter were regional sulfur, which peaked in the warm season, and motor vehicle emissions, which peaked in the cold season. There was insufficient evidence to suggest that exposures to dust, motor vehicle emissions, oil combustion, and regional sulfur were statistically significantly associated with risk of preterm birth in Connecticut. However, insufficient evidence for an association does not imply evidence of no association. Elevated central estimates and previously observed adverse associations with mass concentration motivate the need for further research. Future studies would benefit from study settings with high exposures of a single source to address issues relating to MDL and collinearity, and longitudinal study designs such as that adopted in this study.This was the first study to investigate the effects of PM(1.1 MB) PDFClick here for additional data file.


Studies investigating the relationship between maternal tea drinking and risk of preterm birth have reached inconsistent results.The present study analyzed data from a birth cohort study including 10,179 women who delivered a singleton live birth were conducted in Lanzhou, China between 2010 and 2012.p for trend < 0.01). The relationship between tea drinking and preterm birth is modified by both maternal age (p < 0.05) and gestational weight gain (p < 0.05).Drinking tea , and specifically green  or scented tea , was associated with an increased risk of preterm birth. Drinking tea was associated with both moderate preterm  and spontaneous preterm birth . Risk of preterm birth increased with decreasing age of starting tea drinking  and increasing duration (Despite conflicting findings in the previous literature, we saw a significant association with maternal tea drinking and risk of preterm birth in our cohort. More studies are needed both to confirm this finding and to elucidate the mechanism behind this association. Tea is among the most widely consumed beverage worldwide , 2. Tea The rapid industrialization in China over the past several decades has caused increased deterioration of the environment, which has brought pollution and contamination to tea , 8. HeavPreterm birth (<37 completed weeks of gestation) is the leading cause of child death  and is aA birth cohort study was conducted in the Gansu Provincial Maternity and Child Care Hospital, the largest maternity and child care hospital in Lanzhou, China between 2010 and 2012 [2), normal weight (18.5 kg/m2 ≤ BMI < 24 kg/m2), and overweight and obese (BMI ≥ 24 kg/m2) using the standard of Working Group on Obesity in China [2), 11.5–16 kg (BMI 18.5–23.9 kg/m2), 7–11.5 kg (BMI 24.0–27.9 kg/m2), and 5–9 kg (BMI > 28 kg/m2) [Eligible women were informed of study procedure upon their arrival at the hospital for delivery. After obtaining written consent, trained study interviewers conducted in-person interviews at the hospital using a standardized and structured questionnaire. The majority of women (84 %) were interviewed within three days after delivery, while 16 % of women were interviewed within two days before delivery. Information regarding tea consumption before and during pregnancy was collected. Ever tea drinkers were defined as individuals who drank tea at least three times per week, those who did not fit this description were classified as never tea drinkers . Women win China . Weight BMI 18.5–.9 kg/m2,Preterm birth was defined as delivery before 37 completed weeks of gestation. The gestational age at delivery was calculated in completed weeks from the first day of the last menstrual period. Preterm birth was divided into moderate preterm birth (32–36 weeks of gestation), very preterm birth (28–31 weeks of gestation), and extremely preterm birth (<28 completed weeks of gestation) according to World Health Organization (WHO) classification . To incrχ2 test. Multivariate logistic regression models were used to estimate odds ratios (OR) and 95 % confidence intervals (CI) for the association between tea drinking and preterm birth and its clinical subtypes. Models were adjusted for the following potential confounding factors: maternal age , years of education , employment status during pregnancy (yes or no), monthly income , parity (nulliparous or parous), history of preterm birth (yes or no), hypertension during pregnancy (yes or no), maternal prepregnancy BMI , alcohol consumption (yes or no), and active and/or passive tobacco smoke exposure during pregnancy (yes or no). Additionally, sex of the child, nausea and vomiting during pregancy were included in the models but did not appreciably change the results, therefore they were not included in the final models. All analyses were performed using SAS software, version 9.2 .The relationship between selected characteristcs and tea drinking was analysed using In our study, 7.73 % (787) and 4.38 % (446) women drank tea before and during pregnancy, respectively. 99.6 % of the consumed tea was produced in China. Of the 10,179 singleton live births, 10.01 % (1019) were preterm. Of those 1019 preterm births, 81.75 % (833) were moderate preterm births, 18.25 % (186) were very preterm births, 33.17 % (338) were medically indicated preterm births, and 66.83 % (681) were spontaneous preterm births.Compared to women who never drank tea, those who drank tea were more likely to be older, less educated, parous, drink alcohol and smoke during pregnancy, and have either ≤2000 or >4000 yuan household income Table . Tea driDrinking tea was associated with an increased risk of preterm birth , and for those who began drinking tea before age 20  and GWG (P for interaction = 0.0371). Significantly increased risk of preterm birth was associated with tea drinking among women who were aged 30 years or older , and those who had GWG exceeding the recommendations .We further examined whether maternal age and GWG were effect modifiers Table . SignifiTo the best of our knowledge, this study represents the first to comprehensively examine the associations between tea consumption and preterm birth by various clinical subtypes in a Chinese population. It supports that drinking tea is associated with an increased risk of preterm birth, and that risk varies by preterm birth subtypes.Caffeine, a xanthine alkaloid, is readily available in tea . CaffeinSeveral studies have investigated the association between tea drinking and preterm birth, however, the results were inconsistent , 14–19. In addition to caffeine, heavy metals, pesticides and persistent organic pollutants have also been found in Chinese tea leaves including lead (Pb), chromium (Cr), cadmium (Cd) , 38, perTea consumption is a complicated exposure, as different types of tea have different constitutions and contaminations. For example, black tea contains the highest amount of arsenic; oolong tea contains the highest amount of chromium , while sr = −0.72). It is unclear whether the higher risk of preterm associated with starting drinking tea at younger age is due to longer duration of exposure, immaturity of metabolic organ or enzymes might prolong metabolizing harmful elements [We observed that tea consumption only during pregnancy was not significantly associated with an increased risk of preterm birth, based on a small numbers of exposed cases. Women who only drank tea during pregnancy had short duration of tea consumption, and likely began drinking tea at older ages. Our study found that the risk of preterm birth was higher for those who started drinking tea before age 20 and for those who had longer duration of tea drinking. In this study, those with younger started age had the longer duration of tea drinking (elements . Long teelements .We found that maternal age modified the association between tea consumption and risk of preterm birth. Women who are older would have longer accumulated exposure to environmental pollutants. GWG is correlated with fat retention , increasThere were 1.2 million preterm births in China in 2010 . The estA major strength of our study was the relatively large sample size, which allowed us to explore the associations with tea consumption and preterm birth by various clinical subtypes. In addition, detailed information on tea drinking habits allowed us to comprehensively examine associations with tea drinking by exposure time windows, duration, and type of tea. Birth outcomes and maternal complications during pregnancy were obtained from medical records, which minimized potential disease misclassification. Detailed information on potential confounders such as pre-pregnancy BMI, active and passive smoking, and history of preterm delivery were collected and controlled for in our analysis.Limitations should be considered when interpreting the results. We did not quantify the caffeine concentration from tea drinking, which limited our ability to clarify the relationship between caffeine from tea and preterm birth. Information on exposure was collected through in-person interview before/after delivery; therefore, potential recall bias might exist. However, because in China tea consumption is considered one of healthy lifestyles, and women are not informed to avoid tea consumption during pregnancy, if any recall bias exists, it is likely to be non-differential and would result in an underestimation of the observed association. Lack of quantification of tea consumption prevented us from analyzing more detailed categories of tea drinking. Future studies should collect information on amount of tea consumed per day.In conclusion, this study supports the hypothesis that in a Chinese population drinking tea is associated with an increased risk of preterm birth though the underlying etiological mechanism is still unknown. Additionally, the risk of preterm birth might vary by type of tea consumed, age at first tea consumption, and duration of tea consumption. Future studies are needed to explore the potential mechanism by which certain elements in tea, including potential contaminations, influence risk of preterm birth.


Preterm infants born before the 30th week of pregnancy are especially at risk of perinatal brain damage which is usually a result of cerebral ischemia or an ascending intrauterine infection. Prevention of preterm birth and early intervention given signs of imminent intrauterine infection can reduce the incidence of perinatal cerebral injury. It has been shown that administering magnesium intravenously to women at imminent risk of a preterm birth leads to a significant reduction in the likelihood of the infant developing cerebral palsy and motor skill dysfunction. It has also been demonstrated that delayed clamping of the umbilical cord after birth reduces the rate of brain hemorrhage among preterm infants by up to 50%. In addition, mesenchymal stem cells seem to have significant neuroprotective potential in animal experiments, as they increase the rate of regeneration of the damaged cerebral area. Clinical tests of these types of therapeutic intervention measures appear to be imminent. In the last trimester of pregnancy, the serum concentrations of estradiol and progesterone increase significantly. Preterm infants are removed abruptly from this estradiol and progesterone rich environment. It has been demonstrated in animal experiments that estradiol and progesterone protect the immature brain from hypoxic-ischemic lesions. However, this neuroprotective strategy has unfortunately not yet been subject to sufficient clinical investigation. The prevention of preterm birth represents one of the most significant challenges to the field of obstetrics in the 21st century. Preterm infants born before the 30th week of pregnancy are especially at risk of prenatal mortality and morbidity [PIVH originates in the vascular bed of the germinal matrix, an area of the brain that almost completely disappears as the fetus matures –5 Figur. Blood vPVL most commonly leads to damage of the radiatio occipitalis on the trigonum of the lateral cerebral ventricles and the white matter around the foramen of Monroe , 17 Fig. This inPVL can be caused by both cerebral ischemia and infection. During the genesis and the development of the cerebral vascular bed, vascular watersheds develop in the radiatio occipitalis on the trigonum of the lateral cerebral ventricles and the white matter around the foramen of Monroe –28. The Ascending intrauterine infections can also induce PVL –32. An aIn 2000, Wu and Colford published a meta-analysis of 26 studies on the correlation between chorioamnionitis and infantile cerebral palsy . Their aDue to the fact that PIVH and PVL are complications which especially affect extreme preterm infants, both can be avoided by preventing the baby from being born preterm. Much evidence has shown that patients who have previously experienced a preterm birth or who develop shortening of the cervix to less than 25 mm before the 24th week of pregnancy benefit from the prescription of progesterone –48. The Ascending intrauterine infections, which are often observed in patients at risk of preterm birth, seem to significantly increase the risk of fetal brain damage . BacteriIf a preterm birth seems imminent, the infant's brain should be protected by means of the intravenous application of magnesium –57. With4 for the purposes of fetal neuroprotection [Magnesium is a substance which has been used for decades in the field of obstetrics as a prophylaxis for eclamptic seizures and tocolysis. In a case-control study which included infants weighing less than 1500 grams whose mothers were treated with magnesium  the authotection –78.4 was the combined occurrence of infantile cerebral palsy (of serious or medium severity) or death. No significant difference in the combined risk levels was identified between the therapy and control groups. When the combined results were disaggregated, given similar mortality rates, a significantly lower rate of infantile cerebral palsy was identified among children whose mothers had been treated with magnesium sulfate (1.9 versus 3.5%). Rouse and colleagues concluded from these results that the application of MgSO4 leads to a reduction in the incidence of cerebral palsy among very preterm infants [In August 2008, Rouse and colleagues published the results of the BEAM  study, conducted by the Maternal-Fetal Medicine Units Network . The pri infants .4 trial, one of the five above mentioned prospective studies, at school age. The effects of magnesium on the rate of cerebral palsy and abnormal motor function were no longer evident at this time. Possibly, additional therapeutic interventions in the control group may have improved the health status of these children [In 2009, a Cochrane Review was published on the topic . The fivchildren .4 to prevent one case of infantile cerebral palsy (number needed to treat (NNT)) is dependent on the week of pregnancy in which the birth occurs; it is 52 before the 34th week of pregnancy [The number of women at risk of preterm birth who need to be treated with MgSOregnancy  and 29 bregnancy . In the regnancy .The most commonly reported maternal side-effects of systematic magnesium therapy include thrush, sweating, nausea, vomiting, or skin irritation at the injection site. In addition, a 50% increase in the risk of hypotension and tachycardia was reported (number needed to harm (NNH): 28–30). A higher rate of serious complications such as maternal mortality, cardiac or respiratory arrest, pulmonary edema, respiratory depression, serious postpartal hemorrhage, or increased rate of cesarean sections was not identified.4. A statistically significant effect was first apparent above a moderate dosage of 4 g magnesium sulfate. There was no significant difference between the placebo group and children whose mothers received a lower dosage [In the various studies, differing amounts of magnesium were given to patients. The levels ranged from 4 g to almost 50 g of MgSOr dosage . The totr dosage .The treatment should begin with a bolus injection of 4–6 g within 30 minutes, followed by maintenance doses of 1-2 g/h for 12 hours. The aim of this procedure is to double the magnesium level in the mother's serum. If birth does not occur within 12 hours, the administration of magnesium can be restarted at a later point in time if preterm birth again appears imminent.In an Australian perinatal center, Ow and colleagues investigated the rate of women which can be administered magnesium intravenously under clinical conditions for the purpose of neuroprotection in the event of an imminent preterm birth . Out of The administration of high dosages of magnesium in the event of an imminent preterm birth leads to a reduction in the rate of infantile cerebral palsy and gross motor skill dysfunction , 85.An infant's blood volume at birth can be significantly influenced by the time at which the umbilical cord is clamped . In 1988Several studies published since 1980 have shown that delayed clamping of the umbilical cord can reduce the need for blood and fluid transfusion, as well as the rate of brain hemorrhages and sepsis among preterm infants , 91–94. In order to understand the effect of several other treatment measures which have been predominantly tested in animal experiments, it is important to comprehend the pathophysiological processes occurring during and after an injury. The initial damage caused by an injury is normally the result of insufficient metabolic supply. This leads to a loss of membrane potential, a massive release of excitatory neurotransmitters, and a very strong influx of calcium, which in turn activates proteases, endonucleases, and lipase and thus induces successive cell death . HoweverThis is the point at which therapy with so-called stem cells becomes relevant. Stem cells can be obtained from many different types of tissue. Depending on their origin, they are referred to as neuronal, mesenchymal, or hematopoietic stem cells, and so forth. Mesenchymal stem cells are currently considered to have the most potential for clinical applications. They can be grown easily from bone marrow and from extraembryonic tissue such as the placenta, Wharton's jelly, and umbilical cord stroma –108.Originally it was believed that the applied stem cells multiplied in the damaged region, where they differentiated and replaced the destroyed tissue. However, it quickly became clear that this could not be the way the neuroprotective mechanism worked. The low number of stem cells growing and the insufficient rate at which they differentiate in no way explained the significant neuronal improvements observed. Recent animal experiments have shown that the application of stem cells leads to a significantly improved outcome following hypoxic-ischemic damage –111. ThiVia transmitters, mesenchymal stem cells modulate numerous signaling cascades during apoptosis, neurogenesis, angiogenesis, and synaptogenesis. Increased expressions of fibroblast growth factor-2, epidermal growth factor, glial cell line-derived neurotrophic factor, and sonic hedgehog have been observed . These fβ, and other immunomodulating factors [Postischemic inflammation is most commonly caused by microglia, macrophages of the central nervous system, which originate in bone marrow and migrate to the brain during development, where they then differentiate into microglia. When brain damage occurs, these local microglia are activated, and monocytes from peripheral blood also migrate to the trauma site , 128. Th factors . The app factors , 124. In factors , 132  which contain lipids, proteins, and RNA and which are secreted by various types of cells. Exosomes obtained from mesenchymal stem cells have been shown to reduce the extent of myocardial damage following ischemia in experiments with adult mice .The steroid hormones estradiol and progesterone play a critical role in the growth, differentiation, and function of the reproductive system. However, the peripheral and central nervous system is also affected by these hormones, as shown by the ubiquitous distribution of the relevant receptors –137. EstIn the third trimester of pregnancy, the maternal serum concentration of estradiol and progesterone increases significantly and can reach up to 100 times its original level , 140. Prα und β have been shown to be differently distributed and regulated [α are responsible for the antiapoptotic properties. The activation of the receptor leads to a rapid induction of the insulin-like growth factor 1 (IGF-1) receptor pathway and the associated signal cascade [β-estradiol inhibits the caspase-pathway which plays a key role in apoptosis [The neuroprotective effects of estradiol can be imparted receptor-dependent (genomic and nongenomic) or receptor-independent –149. Theegulated , 135. Boegulated , but difegulated . Two eff cascade . IGF-1 h cascade . In addipoptosis .β, TNF-α, IL 6, COX-2, and ICAM-1 [β2, VCAM-1, CD68, and Iba1 [The neuroprotective effects of progesterone are also mediated by various mechanisms. Progesterone reduces postischemic cellular edema by maintaining the integrity of the blood-brain barrier. Increased expressions of claudin5 and occludin1 have been observed in connection with this process, both of which are proteins that play an important role in the creation of tight junctions. In contrast, it has been shown that the expression of MMP-3 and MMP-9 is reduced. The latter is involved in extracellular tissue degradation , 158. Add ICAM-1 , 149–151and Iba1 –162, facand Iba1 , while tand Iba1 . The admand Iba1 . Howeverand Iba1 .Many more neuroprotective strategies have been investigated in animal experiments. However, a detailed discussion of all of these strategies is outside the scope of this review. We would like to invite interested readers to refer to the relevant literature.


Preeclampsia is associated with fetal growth restriction and low birth weights. Neurotrophins, which mediate neuronal growth and development, are also increased in the placenta and cord blood in preeclampsia. Hence, the aim of this study was to determine whether fetal head growth is altered in preeclampsia, adjusting for growth restriction and other confounding variables.This research included a retrospective cohort study, looking at fetal head circumference at birth, plus a case–control study examining fetal head circumference at mid-gestation. The head circumference at birth analysis consisted of 14,607 pregnancies , delivered between July 2006 and June 2012 at Nepean Hospital, Australia. Head circumference at birth, in addition to other maternal and fetal variables, was sourced from the Nepean Obstetric Database. The head circumference at mid-gestation study consisted of 756 pregnancies , delivered within the same data collection period at Nepean Hospital. Head circumference at mid-gestation was retrieved from an earlier ultrasound scan. Exclusion criteria included >1 fetus, illegal drug use, alcohol consumption, and chronic or gestational hypertension. Generalized linear models were used to analyze fetal head circumference in preeclampsia versus controls, adjusting for confounding variables.Head circumference increased at a greater rate in preeclampsia versus controls, adjusted for gestation, fetal gender, birth weight and length, smoking, maternal BMI, and growth restriction. At mid-gestation, there was no difference in head circumference between preeclampsia and controls.For the first time, this research has suggested increased fetal head growth in preeclampsia, adjusted for confounders. This finding may be explained by altered fetal exposure to neurotrophins in preeclampsia. The long-term neurodevelopmental consequences of preeclampsia remain unclear. Preeclampsia is a leading cause of fetomaternal morbidity and mortality. The exact etiology of preeclampsia remains unsolved, though placental dysfunction is central. Specifically, impaired implantation results in an abnormally shallow placenta, with potential for vascular insufficiency and subsequent fetal growth disturbance , 2. AddiMore than nutrient supply, brain growth is determined by neurotrophins, including brain-derived neurotrophic factor (BDNF). As key mediators of neuronal growth, disorders associated with alterations in BDNF are linked to changes in brain volume , 7. In pHead circumference closely relates to brain volume and is routinely performed on all newborns . The aimThis research included a retrospective cohort study examining fetal head circumference at birth plus a case–control study examining head circumference at mid-gestation. Singleton pregnancies delivered at Nepean Hospital, Australia, between July 1, 2006, and June 30, 2012, were considered eligible. Each included pregnancy had a single normal fetus. Exclusion criteria were chronic or gestational hypertension, alcohol consumption, and illegal drug use. Pregnancy information was sourced from the Nepean Obstetric Database, the information in which is updated and maintained during and after pregnancy and contributes to state-wide data collection.The Ethics Committee of the Sydney West Area Health Service approved this study according to the Declaration of Helsinki.de novo hypertension (systolic BP ≥ 140 mmHg and/or diastolic BP ≥ 90 mmHg) and proteinuria (≥0.3 g/24 h or spot urine protein/creatinine ratio ≥ 30 mg/mmol) after 20 weeks’ gestation, in accordance with recommendations proposed by the International Society for the Study of Hypertension in Pregnancy .U test, as the normality assumption was violated. Categorical characteristics were compared using either Fisher’s exact test or Chi-square test, depending on group size.Continuous maternal and fetal characteristics were compared between groups using Mann–Whitney A multivariate generalized linear model (GLM) was used to model head circumference at birth. Given that mother could be in the database more than once (if the mother had more than one child in the study period), this correlation was accounted for in the model. The effect of preeclampsia was investigated adjusted for gestational age, fetal gender, birth weight, birth length, and fetal growth restriction, as well as the mother’s BMI and whether they smoked during their pregnancy. An interaction between preeclampsia and gestational age was also included in the model to investigate the rate at which head circumference at birth increased.Another GLM was used to model head circumference at mid-gestation. The effect of preeclampsia was adjusted for gestational age at ultrasound, fetal gender, maternal BMI, and smoking status during pregnancy.p-value <0.05 was considered statistically significant. No corrections have been made for multiple comparisons. SAS v9.3  was used for the GLM models.For all analyzes, a two-sided Between July 1, 2006, and June 30, 2012, 22,558 pregnancies were delivered at Nepean Hospital, 753 of which were diagnosed with preeclampsia. Following application of the exclusion criteria, 19,358 pregnancies remained eligible: 588 preeclamptics and 18,770 controls. Enforcement of maternal BMI, gestation, and birth weight cutoffs excluded a further 179 pregnancies. Of the remaining eligible pregnancies, 4,572 were excluded from the model due to missing data, mostly owing to blank data entry for fetal growth restriction and/or maternal BMI. The head circumference at birth GLM therefore consisted of 14,607 pregnancies: 382 preeclamptics and 14,225 controls.The head circumference at mid-gestation analysis commenced with the same 588 preeclamptics which remained following application of the exclusion criteria. Of these, 318 preeclamptics were ineligible because they lacked fetal morphometry data from a mid-gestation scan. The remaining 270 preeclamptics were matched (1:2) with 540 controls, which also contained fetal morphometry data from a mid-gestation scan. Due to missing data, especially maternal BMI, 54 pregnancies were finally excluded. The GLM for head circumference at mid-gestation thus consisted of 756 pregnancies: 248 preeclamptics and 508 controls.In the head circumference at birth population, preeclampsia was associated with an earlier gestation at delivery, increased maternal BMI, nulliparity and smoking, plus decreased birth weight, child length, and head circumference parameters Table .At mid-gestation, the preeclampsia group also demonstrated increased maternal BMI and smoking; however, fetal morphometry was comparable.p-value <0.001). The lines are shown to cross at 36.3 weeks gestation. Before this time, a delivered preeclamptic fetus is estimated to have a smaller fetal head circumference than a control fetus (adjusting for confounders), and the difference between the two fetuses becomes smaller as gestational age increases. Conversely, a preeclamptic fetus born after 36.3 weeks gestation is estimated to have a larger head circumference than a control fetus, and this difference increases with increasing gestational age. Table The GLM for head circumference at birth, which included an interaction between preeclampsia and gestational age, revealed a significantly increased slope in the preeclampsia group versus controls over the gestational ages modeled  = 0.003 higher in preeclampics (−0.08 to −0.09); For the first time, this study has suggested fetal head circumference to increase at a greater rate in preeclampsia, adjusting for growth restriction and other confounding variables. At mid-gestation, fetal head circumference was comparable between preeclampsia and control groups.A major strength of this study is large sample size; however, numerical errors made during data entry became a concern. To address this issue, cutoff criteria for maternal BMI, gestation, and birth weight were applied, the aim being to eliminate incorrect data. The application of cutoffs is unlikely to have introduced significant bias, however, since broad limits were used and relatively few pregnancies were excluded. Indeed, most excluded pregnancies were due to missing data, especially maternal BMI. Since the head circumference at birth model consisted of one measurement for each fetus, caution should be observed in interpreting head circumference changes as changes in velocity. Nonetheless, repeated measures are not feasible on a large sample size and despite its limitations, cross-sectional birth data are generally accepted as the best method of generating growth curves . Finallyin utero.The purpose of this research was to introduce the novel concept of altered fetal head growth in preeclampsia. As mid-gestation head circumference measures were comparable in this study, and birth head circumference measures indicated an increased head growth in preeclampsia, altered neurodevelopment might be a late event. However, the gestation at birth ranges in this study was relatively narrow and different between groups. Hence, further research is required to establish fetal head growth in preeclampsia versus healthy pregnancy, which might include serial measures of head circumference Similar to previous reports, male gender and increased maternal BMI were associated with larger head circumference at birth in this study . Birth wPreeclampsia is associated with altered fetal growth patterns. In particular, it is linked to small for gestational age (SGA) fetuses; a finding frequently explained by decreased placental perfusion , 5, 13. Altered head growth in preeclampsia may be explained by increased fetal exposure to neurotrophins, including BDNF. An essential regulator of normal neurodevelopment, disorders linked to alterations in BDNF demonstrate changes in brain growth , 7, 15. In conclusion, for the first time, this study has suggested fetal head growth to be increased in preeclampsia, adjusted for confounding variables. Further investigation is required, and if these findings are confirmed, the long-term neurodevelopmental consequences of preeclampsia will be the subject of future research efforts.DE contributed to data collection, manuscript preparation, statistics, and final approval. AM contributed to study design, revision of manuscript, and final approval. KM contributed to statistics, review of manuscript, and final approval. MP contributed to study design, revision of manuscript, and final approval. RN contributed to study design, revision of manuscript, and final approval.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


While research indicates that triglycerides in hyperglycaemic women may increase birth weight, little is known about this relationship in euglycemic women. This study examines the relationship between maternal blood lipid status and infant adiposity up to 2 years of age.The Data from 331 mother-child pairs from the ROLO longitudinal birth cohort study was analysed. Maternal dietary intakes were recorded and fasting blood lipids, leptin and HOMA were measured in early and late pregnancy and cord blood. Infant anthropometric measurements and skin-fold thicknesses were recorded at birth, 6 months and 2 years. Correlation and regression analyses were used to explore associations between maternal blood lipid status and infant adiposity.P = 0.03) while cord blood triglycerides were negatively associated with birth weight (P = 0.01). Cord HDL-C was negatively associated with infant weight at 6 months (P = 0.005). No other maternal blood lipids were associated with infant weight or adiposity up to 2 years of age.All maternal blood lipids increased significantly during pregnancy. Maternal dietary fat intake was positively associated with total cholesterol levels in early pregnancy. Late pregnancy triglycerides were positively associated with birth weight (in-utero influences on infant body composition. There may be potential to modulate infant body composition through alteration of maternal diet during pregnancy.Maternal and fetal triglycerides were associated with birth weight and cord HDL-C with weight at 6 months. Thus, maternal lipid concentrations may exert  Obesity in childhood has become prevalent worldwide with the World Health Organisation estimating that over 41 million children aged under 5 years are obese . With suin-utero environment affects fetal development remain largely unknown. There is significant potential for the role of the maternal diet during this time, however with regard to specific macronutrients such as lipids, a dearth of knowledge remains. It is important to identify how specific elements of maternal nutrition interact with the child at birth and in later life to discover the dietary intakes that are associated with the best outcomes for both the mother and child.Maternal nutrition impacts both the mother’s own health and can have a lasting impact on the health of the baby . During It is estimated that glucose accounts for up to 80% of the energy used by the fetus during pregnancy and is the primary fuel for fetal growth . HoweverFor non-pregnant individuals, the recommendations for healthy concentrations of blood lipids are; below 5mmol/L for total cholesterol, below 3mmol/L for low-density lipoprotein cholesterol (LDL-C), below 2mmol/L for triglycerides and above 1mmol/L for high-density lipoprotein cholesterol (HDL-C) . AlthougThere is a scarcity of data on the association between blood lipids during pregnancy and infant outcomes, with the limited knowledge focusing on women with gestational diabetes mellitus (GDM). Triglyceride concentrations in women with hyperglycaemia during pregnancy have been shown to be associated with increased birth weight ,16. WhilP < 0.05) and less glucose intolerance, which was diagnosed using a glucose challenge test measuring serum glucose above 7.8mmol/L  [Analysis was carried out on a subset of 331 mother and child pairs from the ROLO  study. This randomised controlled trial (registration number ISRCTN54392969) was carried out in The National Maternity Hospital, Dublin, Ireland. Ethical approval was granted by the National Maternity Hospital Ethics Committee and maternal written consent was obtained. Eight hundred secundigravida women who did not have gestational diabetes but had previously given birth to a macrosomic baby , and were therefore at increased risk of delivering another macrosomic infant ,20, were2) was calculated. Additional demographic information collected included mothers’ education level and smoking status. Three-day food diaries were completed during each trimester to estimate dietary intake throughout the pregnancy. Dietary fat intake was calculated for each trimester using nutritional analysis software NetWISP version 3.0  which utilises the food composition database from the 6th edition of McCance and Widdowson’s food composition tables.All participants had their height and weight measured by trained healthcare professionals at the first 14-week antenatal visit and body mass index , late pregnancy (28 weeks’ gestation) and a cord blood serum sample was collected at delivery. Serum total cholesterol, HDL cholesterol and triglyceride concentrations were measured using Roche cholesterol oxidase method, lipase/GPO-PAP (glycerol phosphate oxidase-p-aminophenazone) and direct HDL-Roche 3rd generation method respectively, on the cobas c702 module of the Roche Cobas 8000 analyser in accordance with the manufacturer’s instructions . The methods are standardised against the isotope dilution/mass spectrometry (ID/MS) methods. LDL-cholesterol concentration was estimated using the Friedewald equation . Multiant-tests. For parametric data, Pearson correlation was used, and Spearman’s correlation for the non-parametric data to individually measure the correlation between each blood lipid (in early and late pregnancy and cord blood), HOMA, C-peptide and leptin concentration and each of the anthropometric measures of child weight and adiposity . Bivariate associations at a significance of P < 0.1 were considered significant and were further analysed using multiple regression models. Multiple regression models were created separately for each outcome of child weight or adiposity using a forced entry approach to include the following confounders; at birth: mother’s BMI at the first visit, infant gestational age at birth, infant gender, mother’s education status, and mother’s smoking status, and at 6-month and 2-years: gender, age at data collection, mother’s education status and whether the child was breastfed. The final multiple linear regression models that were statistically significant (P < 0.05) were reported as the best predictors of infant weight and adiposity. All statistical analyses were carried out using SPSS  software version 20.0 .Membership of the control or intervention group of the ROLO study had no association with blood lipid levels or infant anthropometry so both groups were analysed together. All variables were assessed for normality using the Shapiro-Wilk test and by visual analysis of histograms. Relationships between the central tendencies were examined using paired sample Table 1). Just over half (54.4%) of the study participants had a BMI over 25 kg/m². Mean infant birth weight was 4.076 kg with 54.7% of infants classified as macrosomic or large-for-gestational-age. Mean birth weight centile was 72.9. Average infant weight was 8.4 kg at 6 months and 13.1 kg at 2 years. Mean maternal daily intake of saturated fat was 72.4g in trimester 1 and 73.0g in trimester 2, mono-unsaturated fat was 28.1g in trimester 1 and 28.4g in trimester 2 then poly-unsaturated fat intakes were 22.9g and 23.1g in trimester 1 and 2 respectively.Mean BMI at 14 weeks’ gestation was 26.4 kg/m², which is in the overweight category (Table 2).Concentrations of total cholesterol, non-HDL cholesterol, HDL-C, LDL-C and triglycerides increased significantly from early pregnancy to late pregnancy (P = 0.02). Likewise, dietary intakes of monounsaturated and polyunsaturated fat were positively associated with total cholesterol at this timepoint . Dietary fat intake was not associated with triglyceride concentrations in early or late pregnancy, however, there was a negative association between intakes of saturated, monounsaturated and polyunsaturated fat in trimester two and triglyceride concentrations in cord blood .Dietary saturated fat intake in trimester one was positively associated with total cholesterol in early pregnancy  (Table 3). Late pregnancy triglycerides were positively associated with birth weight (B = 111.17) while cord blood triglycerides were negatively associated with birth weight (B = -453.75). Based on the adjusted r² value, this model explains 31.8% of the variation in birth weight. The same variables were significant in the model for the outcome of birth weight centile (data not shown). There was no association with HOMA (measured in early or late pregnancy) with triglyceride level however late-pregnancy triglycerides were correlated with cord C-Peptide levels (P = 0.001). When C-Peptide was added to the model as a confounder the association of triglycerides in late pregnancy with birth weight was weakened (P = 0.059).After controlling for confounders , outcomes associated with maternal blood parameters were birth weight, birth weight centile, and weight at 6 months. Birth weight was significantly associated with triglyceride concentrations in late pregnancy and cord as well as leptin concentrations in the cord . When the adjusted regression analysis was conducted separately among normal-weight mothers (BMI <25 kg/m2) and overweight/obese mothers (BMI ≥25 kg/m2), late pregnancy triglycerides were associated with birth weight in overweight and obese mothers only (R² = 0.08 and P = 0.008 versus R² = 0.0003 P = 0.92 in BMI <25 kg/m²) and cord triglycerides were associated with birth weight in mothers with a BMI below 25 only (R² = 0.08 and P = 0.01 versus R² = 0.001 and P = 0.69 in BMI ≥25 kg/m²).When maternal fasting glucose concentrations (in early and late pregnancy) were added into the model as additional confounders for birth weight this attenuated the association of triglycerides in late pregnancy (P = 0.07) however triglycerides in cord blood remained significantly associated (P = 0.02). Upon stratification of these data by maternal BMI at 14 weeks’ gestation, we found that the triglyceride associations differed in overweight or obese mothers compared to normal-weight mothers (Table 4). HDL-C concentration from cord blood was negatively associated with weight at 6 months (P = 0.005) and along with gender, child age, mother’s education level and breastfeeding, this model explained 18.8% of the variance in weight at 6 months . In the final models, no other maternal blood lipids were associated with weight or adiposity up to two years of age.The final multivariate model and third infant outcome that was significantly associated with maternal blood parameters was weight at six months  activity facilitates greater fatty acid placental transfer and could contribute to higher fetal fat accumulation . Animal h weight . They alh weight . FurtherThere is limited knowledge on the relationship between maternal HDL-C concentrations during pregnancy and fetal development and infant growth. While HDL-C is not thought to cross the placenta, it has been shown in animal studies to influence cholesterol metabolism in the placenta which could affect the size of the fetus as a result . We founDietary intakes of fat in early pregnancy were associated with total maternal cholesterol concentrations. Previous research in non-pregnant cohorts has shown that an individual’s intake of fatty acids influences their cholesterol concentrations  and animThis study adds to the current knowledge of associations between maternal blood lipid concentrations during pregnancy and outcomes for the baby. A comprehensive meta-analysis found that women with gestational diabetes have elevated triglyceride concentrations , and resThis study had many strengths in that it was one of few longitudinal studies to have maternal and fetal blood samples at two time points during pregnancy, along with detailed infant anthropometry up to 2 years of age. These data allowed us to investigate the complex relationship between blood markers at each time point and fetal growth in later life. All anthropometric measurements  were collected by trained healthcare professionals rather than based on self-reports. As with all studies there were some limitations. While there was a reasonably large sample size and the cohort was healthy, average maternal BMI was in the overweight category. In addition, these mothers had previously given birth to a macrosomic infant and the offspring were, on average, in the macrosomic range. Analysis was carried out and there was no association between maternal lipid profiles and rates of macrosomia but, that being said, the results presented may not be representative of all pregnant and infant populations.in-utero influence on later infant body composition, and this could be modulated by maternal BMI. Further mechanistic research is required to elucidate the complex relationship between maternal blood lipid concentrations and fetal growth and development. There is a need to establish recommended healthy blood lipid concentrations during pregnancy. With dietary intakes of fat during pregnancy potentially influencing maternal blood lipid profiles, there is the potential to modulate infant body composition by altering the mother’s diet during pregnancy.In the present cohort, maternal blood lipid concentrations in late pregnancy and in cord blood were associated with offspring anthropometry. Maternal triglyceride concentrations were associated with birth weight and HDL-C in cord blood was negatively associated with infant weight at 6 months of age. These results suggest that maternal lipid concentrations may exert an


The cyst in mixed dentition stage cause expansion of buccal cortex, displacement of teeth and may present as case of infection. The cyst in periradicular region of deciduous molar are of frequent occurrence. The differential diagnosis of this lesion is radicular cysts of deciduous molar: developmental or infammatory dentigerous cyst of corresponding unerupted premolar. After going through the available literature of radicular cyst of deciduous molars and dentigerous cysts of developing premolars in mixed dentition we studied the five cases of cyst in periradicular region of deciduous molars in mixed dentition retrospectively for the diagnostic dilemma of radicular cyst verses dentigerous cyst. In conclusion, we can set some criteria for the diagnosis which is particularly important for treatment and for conservation of developing premolar.How to cite this article: Manekar VS, Chavan A, Wadde K, Dewalwar V. Cysts in Periradicular Region of Deciduous Molars in Mixed Dentition: Retrospective Study of Five Cases. Int J Clin Pediatr Dent 2014;7(3):229-235. This is particularly important for treatment, prevention of localized malocclusion and conservation of premolar toothbud. Aim – Retrospective study of cases of cyst in periradicular region of deciduous molars in mixed dentition. Objective – To set criteria for diagnosis of radicular cyst vs dentigerous cyst. Inclusive criteria-the cases diagnosed as cyst in posterior jaw of mixed dentition with radiolucency in periradicular region of deciduous molars with corresponding premolar bud in lesion. Cases reported to Department of Oral and Maxillofacial Surgery, at Government Dental College and Hospital, Mumbai were studied. There were five patients with diagnosis of infected cyst with deciduous molar region in mixed dentition; they were comparatively studied for their clinical presentation, radio-graphic presentation, and differential diagnosis, outcome of treatment and for criteria of definitive diagnosis of radicular cyst of deciduous molar or dentigerous cyst of developing premolar.The cyst in periradicular region of deciduous molars in mixed dentition present with the symptoms of swelling, pus discharge, pain and over retained teeth. The expansion of buccal cortex and displacement of corresponding developing premolar is a common presentation. These cysts can be radicular cysts of deciduous molars or; developmental or infammatory dentigerous cysts of corresponding premolar or supernumerary tooth bud. In case of carious or endodontically treated deciduous molar, there is possibility of radicular cyst of deciduous molars or inflammatory dentigerous cyst of corresponding premolars. After going through the available literature of radicular cyst of deciduous molars and dentigerous cysts of premolars we studied the five cases of cyst in periradicular region of deciduous molars in mixed dentition treated in our department for the diagnostic dilemma of radicular cyst Case 3 is cystic lesion with carious 85, around its peri-apical and furcation; 45 tooth engulfied in well corticated lesion . MarsupiCase 5 is cystic lesion with endodontically treated, over retained 85. Out of five cases reported here three were labeled as dentigerous cyst and two were labeled as radicular cyst. The histopathological report of all cases is infected dental cyst. Hence, histopathology was not useful for diagnosis. The diagnosis is done on clinical, radiological and surgical correlation. 1 Cysts associated with deciduous molars are usually in their periapical region and involve the developing premolar teeth and the patients are bellow 12 to 13 years that is before eruption of premolars or in later age group with over retained molars and delayed eruption of premolars. Shear (1994) has estimated that about 9% of dentigerous and 1 % of radicu lar cysts occur in the first decade of life, while according to Donath (1985) about 4% of dentiger-ous and less than 1% of radicular cysts appear in this life period. In the study of Lustmann and Shear (1985), of radicular cysts with deciduous teeth in 23 cases, the mandible was affected more frequently than maxilla, and the deciduous molars were the teeth most often involved. In nine cases buccal expansion was noticed and in eight cases permanent buds were displaced. The caries was the most common etiological factor.1 According to Shear, Radicular cysts of deciduous teeth appear to be rare. Radicular radiolucency related to deciduous teeth tend to be neglected and probably resolve after removal of offending tooth.1Radicular cyst is one which arises from the epithelial residues in the periodontal ligament as a result of infam-mation. The infammation usually follows the death of the dental pulp and cysts arising in this way are found most commonly at the apices of the involved teeth.2 Periradicular cysts that have been described in association with pulpotomized primary molars seem to show specific clinical features: large size, rapid growth, buccal expansion, and displacement of succedaneous teeth.3 Dentigerous cyst or follicular cyst is an odontogenic cyst associated with the crown of an impacted, embedded, unerupted or developing tooth. The cyst which encloses the crown of an unerupted tooth is attached to the cervical region of the tooth. It is the second most common type of odontogenic cysts accounting for 14 to 24% of all jaw cysts.4 The relationship of cyst development to the mandibular second primary molars cariously affected or pulpotomized has been stressed.5 There are two possible explanations: (1) that the mandibular molars have a greater susceptibility to caries and, consequently, are more frequently treated; and (2) that a mandibular second primary molar is more closely associated with its successor's follicle; such association can more easily facilitate the spread of infammation in comparison with other primary teeth.6 Shaw et al noted that distinguishing between dentigerous and radicular cysts on histologic grounds is difficult but that a histologic examination may determine whether the lesion is chronic or acute in nature.6 In our experience all five cases shows histological features of infected dental cyst .9 In the periradicular region of pulpotomized deciduous molars, besides the dentigerous cyst, a glandular odontogenic cyst can develop.2Periapical cysts in the primary dentition are located around the roots and in the inter-radicular area; this is in contrast to the periapical location seen in permanent teeth. Infammation and prolonged irritation are apparent causes of, and the trigger for, proliferation of the epithelial rests. This explains why cysts are found in conjunction with necrotic or pulp-treated teeth in the primary dentition.tal cyst  hence hi10 several differences can been found between the radiological features of periradicular cysts and dentigerous cysts: periradicular cysts are (1) frequently unilocular, although occasionally they are multilocular; (2) they are well-defined radiolucencies, located both periradicularly and inter-radicularly; and (3) the pericoronal space of the underlining permanent tooth appears normal, usually with an intervening total or partially distinct cortical layer of bone. On the contrary, dentigerous cysts: (1)generally have no distinct boundaries between the roots of the primary tooth, and the crown of the underlying succeeding tooth; (2) a single, unilocular, well-defined, radiolucent area encloses each kind of cyst; and (3) a radiolucent area that embraces the permanent tooth bud, ill-defined lesion borders, and arrest of the root development are pathognomonic signs of damaged tooth germs.As mentioned by Asian-Gonzélez et al11 state that, treatment modalities range from enucleation to marsupialization. It is imperative that utilization of a conservative approach to treatment of dentigerous cysts that does not involve, sacrifice of the unerupted permanent tooth whenever possible. However, complete eradication of the cyst should never be compromized for saving the permanent tooth bud. In cases in which the permanent tooth is severely damaged, and hopelessly displaced, complete enucleaton of the cyst to include the permanent tooth bud has been recommended .12Ziccardi et al (1997)Criteria of differentiation as from literature and our study are- radicular cyst- carious or endodonticaly treated deciduous molar; more chances of signs of infection, radiolucency around the roots, involving the furcation as well, complete premolar tooth bud looks engulfied in the lesion on OPG.No carious lesion with deciduous molar; severe displacement of involved developing premolar as well as adjacent premolar; radiographically and after excision lesion looks typically around premolar crown, even if it is displaced.Expansion of buccal cortex, case presenting as infection, displacement of teeth; aspiration of straw colored fuid unless infected, resorption of roots of deciduous molars, well defined cystic radiolucency with cortication.vs dentigerous cyst of corresponding premolar. This is particularly important for treatment purpose, for preservation of developing premolar and the localized malocclusion developed due to delayed development. We studied the five cases retrospectively for this diagnostic dilemma, and we could set some criteria – three cases of dentigerous cysts and two cases of radicular cyst. Though the sample size of this study is very small, the distinction noted in cases of radicular cyst of deciduous cyst and dentigerous cyst of developing premolar are enough to laid down the criteria for their diagnosis. Although, the studies with larger sample sizes are recommended.The differential diagnosis of periradicular cyst of deciduous molars with carious lesion or endodontically treated can be radicular cyst; or Infammatory denti-gerous cyst of corresponding premolar. The periradi-cular cysts of intact deciduous molars are developmental dentigerous cysts of corresponding premolars. It is a diagnostic dilemma of radicular cyst of deciduous molar


The ability of HDL to support macrophage cholesterol efflux is an integral part of its atheroprotective action. Augmenting this ability, especially when HDL cholesterol efflux capacity from macrophages is poor, represents a promising therapeutic strategy. One approach to enhancing macrophage cholesterol efflux is infusing blood with HDL mimics. Previously, we reported the synthesis of a functional mimic of HDL (fmHDL) that consists of a gold nanoparticle template, a phospholipid bilayer, and apo A-I. In this work, we characterize the ability of fmHDL to support the well-established pathways of cellular cholesterol efflux from model cell lines and primary macrophages. fmHDL received cell cholesterol by unmediated (aqueous) and ABCG1- and scavenger receptor class B type I (SR-BI)-mediated diffusion. Furthermore, the fmHDL holoparticle accepted cholesterol and phospholipid by the ABCA1 pathway. These results demonstrate that fmHDL supports all the cholesterol efflux pathways available to native HDL and thus, represents a promising infusible therapeutic for enhancing macrophage cholesterol efflux. fmHDL accepts cholesterol from cells by all known pathways of cholesterol efflux: unmediated, ABCG1- and SR-BI-mediated diffusion, and through ABCA1. HDL performs a critical role in the pathogenesis of coronary artery disease (CAD) and acute coronary syndrome . The invGiven its involvement in CAD pathogenesis, HDL-CEC is an attractive target for an antiatherogenic therapeutic intervention with the goal of increasing the transport of cholesterol from macrophages to the liver –15. CholPreviously, we reported the synthesis of a nanostructure designed to mimic the ability of HDL to accept cellular cholesterol , 23. Thi3 (1.125 g/ml < d < 1.21 g/ml) were isolated by sequential ultracentrifugation from a pool of normolipidemic frozen human plasma as described by Havel, Eder, and Bragdon (3H]apo A-I (∼1 μCi/mg) was prepared by reductive methylation of apo A-I with [3H]formaldehyde  in the presence of sodium cyanoborohydride followed by dialysis in 0.1 M NaCl (3 (−1cm−1 for apo A-I.HDL (1.065 g/ml < d < 1.21 g/ml) and HDL Bragdon , stored  Bragdon . Traut’s Bragdon , and thu1 M NaCl . Protein NaCl (3  and 1.13sn-glycero-3-phosphoethanolamine-N-[3-(2-pyridyldithio)propionate]  (PDP PE) and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) (both from Avanti Polar Lipids) were dissolved in ethanol before each particle preparation. PDP PE and DPPC were sequentially added to the mixture, both in 250-fold molar excess relative to the starting amount of Au NPs. The mixture was returned to the shaker and incubated overnight (∼12 h). For fmHDL without apo A-I [fmHDL (no apo A-I)], the synthesis was performed by the same procedure except the addition of apo A-I was omitted.fmHDL was synthesized using citrate-stabilized colloidal Au NPs (Ted Pella). Transmission electron microscopy (TEM) analysis of the Au NPs sold as “5 nm in diameter” revealed that the diameter of these nanoparticles was actually 6 ± 1 nm . A 5-foli tangential flow filtration system (Spectrum Laboratories) was employed to purify fmHDL by diafiltration, a combination of filtration and dialysis in which impurities are removed as the sample passes over a semipermeable membrane  on a Cary 5000 UV-Vis-NIR spectrophotometer (Agilent Technologies), as Au colloid absorbs light strongly at ∼520 nm due to the surface plasmon resonance  to obtain apo A-I-SH. The molar concentrations of apo A-I-Alexa 488 and apo A-I-SH-Alexa 488 were determined by absorbance as recommended by the dye manufacturer. Standard curves of apolipoprotein concentration versus Alexa 488 fluorescence were constructed separately for apo A-I-Alexa 488 and apo A-I-SH-Alexa 488 by serial dilution of the dye-labeled apolipoproteins with PBS/80 mM KCN (Sigma-Aldrich)/0.01% Tween 20 (Sigma-Aldrich) in a 96-well plate. Fluorescence was measured at 520 nm (490 nm excitation) using a BioTek Synergy H4 multi-mode microplate reader. fmHDL particles were synthesized with the same apo A-I-Alexa 488 (two different preparations) or apo A-I-SH-Alexa 488 (three different preparations) and purified by diafiltration (see above). fmHDL concentrations were determined based on Au NP absorbance at 524 nm. Aliquots of the fmHDL samples were diluted with water and PBS/80 mM KCN/0.01% Tween 20  to 10 nM fmHDL in a 96-well plate and read with the plate reader. KCN and Tween 20 were added to dissolve the Au NPs that would otherwise quench fluorescence and to solubilize the proteins and phospholipids, respectively. Fluorescence measurements were converted to molar concentrations of apo A-I-Alexa 488 and apo A-I-SH-Alexa 488 using the standard curves.The strong absorbance of UV and visible light by Au NPs and of UV light by the PDP PE 2-pyridyl group rendered the common absorbance-based protein quantification methods unreliable and precluded direct determination of apo A-I or apo A-I-SH concentration in each fmHDL sample. A fluorescence-based method was used to quantitate apolipoprotein content in representative fmHDL (apo A-I) and fmHDL (apo A-I-SH) samples, as previously described . Apo A-Im/z 270:284).The palmitate content of fmHDL preparations was measured at the Metabolic Tracer Resource . Fifty microliter subsamples of 4.71 μM fmHDL preparations were combined with heptadecanoic acid  as an internal standard  and 200 μl of 0.3 N KOH in ethanol and incubated at 70°C for 30 min to saponify phospholipid. Palmitic acid standards ranging from 0 to 200 μM were made using a gravimetrically prepared stock solution of palmitic acid . Fifty microliters of each standard were combined with heptadecanoic acid internal standard . Following saponification of fmHDL samples, fatty acids in samples and standards were esterified with 14% boron trifluoride in methanol and analyzed on an Agilent 7890A/5975C series GC/MSD (Agilent Technologies) using electron ionization. Palmitate peak abundance was expressed relative to the internal standard and quantified using a standard curve ] × dilution factor/32,430 M−1, as recommended by the dye manufacturer) versus Alexa 488 fluorescence. The moles of apolipoprotein were divided by the moles of Au NPs to calculate the average number of apo A-I and apo A-I-SH molecules per Au NP and thus, per fmHDL. In a different approach, fmHDL (apo A-I) was synthesized with [3H]apo A-I. The molar concentration of Au NP was calculated as above, whereas the molar content of apo A-I was derived from a standard curve of apo A-I concentration  versus [3H]apo A-I counts per minute values. To determine the average number of phospholipid molecules per fmHDL, fmHDL was prepared normally, the Au NP concentration was measured as described above, and the molar concentration of palmitate in fmHDL was determined using gas chromatography-mass spectrometry (see above). Because both PDP PE and DPPC contain two palmitoyl acyl chains, the palmitate molar concentration was divided by two to calculate the molar concentration of phospholipid. The molar concentration of phospholipid was divided by the molar concentration of Au NP to determine the average number of phospholipid molecules per fmHDL. In a radiolabel-based approach, fmHDL (apo A-I) was made with L-α-dipalmitoyl,[choline-methyl-3H]phosphatidylcholine (PerkinElmer), and the molar concentration of DPPC in the fmHDL samples was calculated from a standard curve of [3H]DPPC concentration versus [3H]DPPC counts per minute. All stoichiometric data are provided in The average number of apo A-I or apo A-I-SH molecules per fmHDL particle was determined using fmHDL preparations made with apo A-I-Alexa 488 or apo A-I-SH-Alexa 488 (see above). The molar concentration of Au NPs in the preparations was calculated by the formula: A3) (Thermo Scientific) was added to the final concentration of 2.5 mM, and cross-linking reactions proceeded for 30 min at room temperature. To generate an apo A-I oligomer ladder, lipid-free apo A-I was dialyzed in 10 mM sodium phosphate buffer (pH 7.4), diluted with the same buffer to 0.5 mg/ml, and cross-linked with 0.25 mM BS3 at room temperature for 30 min. The 0.5 M Tris base  was added to stop the reactions. Cross-linked products were resolved on 4–20% Tris-glycine SDS-PAGE gels and visualized via immunoblotting with a goat polyclonal anti-apo A-I antibody .fmHDL samples were diluted to 50 μg/ml apolipoprotein with water/10× sodium phosphate buffer . Bis[sulfosuccinimidyl] suberate (BSA pair of tweezers was used to hold a 200 mesh copper grid covered with a carbon film (Electron Microscopy Sciences), while a 5 μl drop of 500 nM fmHDL was pipetted onto the grid’s carbon side. The drop was allowed to adsorb to the grid for 15 min. Then the excess solution was wicked away with filter paper, and the grid was left to dry for five more minutes. A 40 μl drop of 2% (w/w) uranyl acetate (UA) in water was pipetted onto a small piece of parafilm. The grid was floated on the UA drop with the sample side down for 7 min and then placed on a piece of filter paper with the sample side up to dry. The UA solution was shielded from light to minimize stain degradation. TEM images were taken with a FEI Tecnai Spirit G2 transmission electron microscope operating at 120 kV. Proteins and phospholipids have low electron density and require staining for TEM imaging. In UA-stained samples, apolipoproteins and phospholipids are visible as white rings around the electron dense Au NP.Size and zeta potential were determined by DLS with a Zetasizer Nano ZS . Measurements were taken with the following parameters: 50–100 nM of fmHDL in water, material refractive index of 0.30, and absorbance of 0.234. At least five measurements were averaged for each sample, and at least three different fmHDL preparations were measured. The number function was used to determine the hydrodynamic diameter.For circular dichroism (CD), 1.5 μM of apo A-I or 500 nM of fmHDL (three different preparations) were used. Background was subtracted for each measurement. The background for apo A-I was water (three samples) or 1× PBS with 4 mM EDTA at pH 8.0 (one sample). The solvents did not affect the apo A-I spectra. fmHDL (no apo A-I) was used as the background for apo A-I-containing fmHDL to subtract the CD signal from the Au NPs and phospholipids. Measurements were done with a Jasco J-815 CD spectrophotometer. Three different databases  were used to analyze the CD spectra, and the average of the results from the three databases is reported as the percentage of residues having a particular secondary structure.2. Primary resident peritoneal mouse macrophages were obtained from 2- to 4-month-old all male or all female C57BL/6 mice -ABCA1 and BHK-ABCG1 cells were kindly provided by the late Dr. Jack Oram . BHK-SR-L/6 mice . Periton6 and apo A-I-SH micrograms per milliliter = [fmHDL] × 4 × 28,080/106, where [fmHDL] is the concentration of fmHDL calculated from A524 (see above), 3.3 and 4 are the number of apo A-I and apo A-I-SH, respectively, molecules per fmHDL  and 0.118 × 4 × 28,080/106 = 13.3 μg/ml in terms of apo A-I-SH in fmHDL (apo A-I-SH). Four-fold higher amounts (0.472 μM) were used in the SR-BI-mediated cholesterol efflux assays in BHK-SR-BI and Fu5AH cells, and 0.236 μM (2×) amounts were used for cholesterol efflux assays in mouse macrophages. All protein in fmHDL was apolipoprotein. The concentration of HDL3 in terms of the total protein content was matched to the apo A-I concentration of fmHDL (apo A-I).As mentioned above, protein measurement methods based on absorbance were not suitable for apo A-I quantification in fmHDL preparations. The apolipoprotein content of fmHDL was calculated by the equations: apo A-I micrograms per milliliter = [fmHDL] × 3.3 × 28,080/103H(N)]cholesterol (PerkinElmer), and the cells were incubated for 24 h to label the intracellular pools of cholesterol. On day 3, the medium was changed to DMEM supplemented with 0.2% BSA , 50 μg/ml gentamicin, and either 10 nM mifepristone (Sigma-Aldrich) or vehicle, and the cells were incubated overnight for 16–20 h. On day 4, the medium was changed to DMEM supplemented with 50 μg/ml gentamicin and either 10 nM mifepristone or vehicle, with or without a cholesterol acceptor , and the cells were incubated for 4 h (2 h in the case of BHK-SR-BI). fmHDL suspended in water comprised 10% of the cholesterol efflux medium. To account for any effects of medium dilution, water was added to 10% to acceptor-free and HDL3- or apo A-I-containing media. The latter two cholesterol acceptors were suspended/dissolved in TBS. To account for any effects of TBS on cholesterol efflux, an equivalent amount of TBS was added to the fmHDL and acceptor-free media. At the end of the incubation period, cell media were collected and filtered through 96-well filter plates (EMD Millipore). A 100 μl aliquot of each sample was read in a scintillation counter. Cell lipids were extracted with hexane-isopropanol , the solvent was evaporated, and the lipids were read in a scintillation counter. [3H] counts in the medium were multiplied by five, divided by the sum of [3H] counts in the medium and in the cells, and multiplied by 100 to calculate the percentage of total cellular cholesterol released to the cholesterol acceptors.BHK-ABCA1, BHK-ABCG1, and BHK-SR-BI cells were seeded at 1/10 dilution from confluent flasks into 24-well plates in DMEM supplemented with 10% FBS and 50 μg/ml gentamicin (500 μl of medium/well) and allowed to attach overnight. On day 2, the medium was changed to DMEM supplemented with 2.5% FBS, 50 μg/ml gentamicin, and 0.5–1.0 μCi/ml cholesterol in DMEM supplemented with 2.5% FBS and 50 μg/ml gentamicin overnight. On day 3, the cells were exposed to 200 μg/ml LDL in DMEM supplemented with 0.2% BSA and 50 μg/ml gentamicin overnight. On day 4, the cells were treated with probucol (Sigma-Aldrich), BLT-1, or vehicles for 1–1.5 h and then exposed to fmHDL in the presence of the same inhibitors for 4 h, in both cases in DMEM supplemented with 50 μg/ml gentamicin. Cell media and cells were analyzed as described above. To introduce probucol into the medium, DMEM supplemented with 50 μg/ml gentamicin in a 15 ml conical tube was heated to 37°C in a water bath. Probucol (20 mM) in ethanol was pipetted into the preheated medium while it was vigorously vortexed.Primary macrophages, seeded as outlined above, were labeled with 0.5 μCi/ml cholesterol (PerkinElmer) alone or 0.06 μCi/ml [4-14C]cholesterol and 1.3 μCi/ml [methyl-3H]choline chloride (PerkinElmer) in DMEM supplemented with 2.5% FBS and 50 μg/ml gentamicin overnight. On day 3, the cells were treated with 10 nM mifepristone or vehicle for 16–20 h in DMEM supplemented with 0.2% BSA (fraction V) and 50 μg/ml gentamicin. Then the cells were exposed to fmHDL (apo A-I), fmHDL ([3H]apo A-I), apo A-I, or acceptor-free medium in DMEM supplemented with 50 μg/ml gentamicin for 8 h. fmHDL ([3H]apo A-I) was also added to DMEM supplemented with 50 μg/ml gentamicin in T75 flasks that did not contain cells (no cells control). Cell media were collected, filtered through 0.45 μm polyvinylidene difluoride (PVDF) membrane filter units (EMD Millipore), reduced in volume 20-fold from 30 to 1.5 ml using Amicon Ultracel-10K centrifugal filters (EMD Millipore), and stored at 4°C for further analysis. A 1 ml aliquot of the 20-fold concentrated cell medium was resolved into 1 ml fractions on a calibrated HiLoad 16/60 Superdex 200 gel filtration column . TBS (pH 7.4) was the mobile phase. Five hundred microliters of each fraction were transferred to a cuvette to measure A523. The remainder was combined with 5 ml of ScintiVerse BD cocktail (Fisher Scientific) and read in a scintillation counter. [3H] and [14C] counts were adjusted for the energy emission spectra overlap. The gel filtration column was washed between runs with 30% isopropanol and 1 M NaOH, as recommended by the column manufacturer. The following standards (Sigma-Aldrich) were used to calibrate the column: cytidine, Vt; thyroglobulin, 17.0 nm; apoferritin, 12.2 nm; lactic dehydrogenase, 8.16 nm; BSA, 7.1 nm; carbonic anhydrase, 4.4 nm; blue dextran, Vo.BHK-ABCA1 cells (three T75 flasks per cholesterol acceptor) were plated at 1/10 dilution from a confluent culture and allowed to attach overnight in DMEM supplemented with 10% FBS and 50 μg/ml gentamicin. On day 2, the cells were labeled with 0.06 μCi/ml apo A-I, and the average number of apo A-I molecules per particle was determined using the [3H]apo A-I specific activity. This method showed that fmHDL (apo A-I) contained 3.3 ± 0.2 molecules of apo A-I per particle. Apolipoproteins on fmHDL were also cross-linked with BS3, an amine-to-amine protein cross-linker, and then resolved on an SDS PAGE gel and visualized by an anti-apo A-I antibody  contained 190 ± 20 molecules of DPPC per fmHDL. The larger number of DPPC than PDP PE molecules per particle, 190 versus 170 (360 minus 190), is in line with our working model of fmHDL, which suggests that DPPC preferentially forms the more voluminous outer monolayer, while PDP PE preferentially comprises the less voluminous inner monolayer  and fmHDL (apo A-I-SH). CD was used to characterize the secondary structure of apolipoprotein in lipid-free and fmHDL-associated forms .Mature spherical HDL acquires macrophage cholesterol by unmediated and ABCG1- and SR-BI-facilitated diffusion; it can also release lipid-poor apo A-I, which accepts cholesterol via ABCA1 , 16, 17.3. fmHDL samples were added to efflux media to obtain the final particle concentration of 0.118 μM. Because of the difference in the number of apo A-I molecules per particle .BHK-ABCG1 cells treated with mifepristone or vehicle were incubated with one of the three types of fmHDL or with HDLparticle , the finparticle , 34, 40. amounts . Thus, H in HDL3  versus 3 in HDL3 . HDL3 wa3. The percent efflux values were 3.4 ± 0.5, 2.8 ± 0.5, 1.5 ± 0.2, and 1.7 ± 0.6 for fmHDL (apo A-I), fmHDL (apo A-I-SH), fmHDL (no apo A-I), and HDL3, respectively (mean adjusted for efflux to medium without a cholesterol acceptor ± SD). All of the cholesterol acceptors also acquired cholesterol by ABCG1-facilitated diffusion , but the differences were not statistically significant. Thus, relative to HDL3, all fmHDL particles ensure robust cellular cholesterol efflux by unmediated and ABCG1-facilitated diffusion.All of the acceptors acquired cholesterol by unmediated diffusion  reflected a balance between the need for higher acceptor levels and the technical ability to synthesize fmHDL at high concentrations. All of the acceptors robustly supported unmediated cholesterol diffusion from BHK-SR-BI cells, as was observed for BHK-ABCG1 cells  and fmHDL (apo A-I-SH), respectively  was observed, suggesting that apo A-I is essential for the ABCA1 pathway to function, as is expected  eluted as a broad peak centered at the 14.1 nm mark, a size value similar to the one obtained by DLS  took place in the absence of ABCA1. For fmHDL exposed to ABCA1-expressing cells, the A523 peak overlapped with an enlarged [14H]cholesterol peak and a prominent [3H]choline-phospholipid peak, indicating that ABCA1 supported cholesterol and phospholipid efflux to fmHDL (apo A-I). However, the center of the fmHDL phospholipid peak (12.5 nm fraction) eluted after the center of the fmHDL A523 peak, but well before the peak for discoidal HDL. This elution pattern suggests that smaller  fmHDL (apo A-I) are more efficient recipients of cellular phospholipid by the ABCA1 pathway than the larger  species.ABCA1-mediated cholesterol efflux to fmHDL was further investigated by gel filtration in conjunction with cell lipid radiolabeling . BHK-ABCd by DLS . For fmH3H]apo A-I was used to determine whether ABCA1-mediated efflux to fmHDL (apo A-I) was due to apo A-I dissociation from fmHDL, followed by ABCA1-dependent lipidation of the lipid-poor apolipoprotein. BHK-ABCA1 cells were labeled with [14C]cholesterol, treated with vehicle or mifepristone, and exposed to fmHDL ([3H]apo A-I) or medium without a cholesterol acceptor (3H]apo A-I) was incubated in tissue culture flasks without cells to produce a no-cells control. BHK-ABCA1 cells concurrently form two sizes of discoidal HDL: 10–12 nm and 7–8 nm  than the corresponding fractions of the vehicle-treated and no-cells controls. The source of this additional [3H]apo A-I in the 10–12 nm region was not apparent. The [3H]apo A-I traces for fmHDL (apo A-I) exposed to vehicle-treated cells and fmHDL (apo A-I) exposed ABCA-expressing cells tracked each other closely in the 17.7–12.5 nm size region that includes the fmHDL (apo A-I) A523 and cholesterol peaks . [3. [33H]ap]apo A-I . The 10–ol peaks . This im3 . The magnitudes of cholesterol efflux to all three acceptors did not differ significantly, indicating that fmHDL (apo A-I) functioned just as well as native HDL  to double . There wMacrophage HDL-C flux, the net transport of macrophage cholesterol in HDL to the liver for secretion, is a critical atheroprotective factor and a promising target for therapeutic intervention . One appGold NPs are a flexible platform on which to assemble requisite components. fmHDL contains phospholipid and apo A-I, but other elements can also be readily added . In termCells release cholesterol by four well-documented processes: unmediated diffusion and ABCG1-, SR-BI-, and ABCA1-mediated pathways , 16. UnmfmHDL (apo A-I) also supported the ABCA1 pathway –7. ABCA1Previous ex vivo and in vivo investigations have shown that cholesterol-normal and cholesterol-loaded macrophages employ the cellular cholesterol efflux pathways differently , 53. An A potential advantage of fmHDL is that it is suitable for further functionalization. One possibility is the inclusion of macrophage-specific markers, which would impart fmHDL with enhanced targeting properties . In addi