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Ovarian cancer (OvCa), while accounting for only 3% of all women’s cancer, is the fifth leading cause of cancer death among women. One of the most significant obstacles to successful OvCa treatment is chemoresistance. The current lack of understanding of the driving mechanisms underlying chemoresistance hinders the development of effective therapeutics against this obstacle. Adipocytes are key components of the OvCa microenvironment and have been shown to be involved in OvCa cell proliferation, however, little is known about their impact on OvCa chemoresistance. In the current study, we found that adipocytes, of both subcutaneous and visceral origin, secrete factors that enhance the resistance of OvCa cells against chemotherapeutic drugs by activating the Akt pathway. Importantly, we have demonstrated that secreted lipids mediate adipocyte-induced chemoresistance. Through a comprehensive lipidomic analysis, we have identified this chemo-protective lipid mediator as arachidonic acid (AA). AA acts on OvCa cells directly, not through its downstream derivatives such as prostaglandins, to activate Akt and inhibit cisplatin-induced apoptosis. Taken together, our study has identified adipocytes and their secreted AA as important mediators of OvCa chemoresistance. Strategies that block the production of AA from adipocytes or block its anti-apoptotic function may potentially inhibit chemoresistance in OvCa patients.

Objectives Nasal potential difference (NPD) measurement is part of the diagnostic criteria for cystic fibrosis (CF) and now used routinely as an endpoint in clinical trials of correcting the basic defect in CF. Intestinal current measurement (ICM), measured ex vivo on a rectal biopsy, has been used to study cystic fibrosis transmembrane conductance regulator (CFTR) function but has not been compared to NPD in the same subject in adults and children. The aim of the study is to evaluate the potential usefulness of ICM as a marker of CFTR function for treatment studies compared NPD in patients with CF and in healthy control subjects. Methods ICM and NPD were performed on healthy controls and patients with CF. The healthy adults were individuals undergoing routine screening colonoscopy at the Beth Israel Deaconess Medical Center. The healthy children were undergoing colonoscopy for suspicion of inflammation in Hadassah Hebrew University Medical Center. The CF adults were recruited from Boston Children's Hospital CF Center and CF Center Worcester Mass, the children with CF from Hadassah CF Center. Results ICM measurements in healthy control subjects (n = 16) demonstrated a mean (±SE) carbachol response of 16.0 (2.2) μA/cm, histamine response of 13.2 (2.1) μA/cm and a forskolin response of 6.3 (2.0) μA/cm. Basal NPD of -15.9 (1.9) and response to Cl free + isoproterenol of -13.8 (2.0). These responses were inverted in CF subjects (n = 12) for ICM parameters with carbachol response of -3.0 (0.5) μA/cm, histamine -1.0 (0.8) μA/cm and a forskolin response of 0.5 (0.3) and also for NPD parameters; basal NPD of -42.2 (4.3) and response to Cl free + isoproterenol of 4.3 (0.7). Pearson correlation test showed the comparability of ICM and NPD in assessing CFTR function. Conclusions ICM is equivalent to NPD in the ability to distinguish patients with CF from controls and could be used as surrogate markers of CFTR activity in treatment protocols.

Aliment Pharmacol Ther 2012; 35: 255–265 Summary Background Primary sclerosing cholangitis (PSC) is characterised by progressive inflammatory and fibrotic destruction of the biliary ducts. There are no effective medical therapies and presently high dose ursodeoxycholic acid is no longer recommended due to significant adverse events in a recent clinical trial. Cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction is associated with PSC in both children and adults. Since CFTR dysfunction leads to altered fatty acid metabolism, specifically reduced docosahexaenoic acid (DHA), we hypothesised that DHA supplementation might be an effective therapy for patients with PSC. Aim To determine the safety and efficacy of oral DHA supplementation for the treatment of PSC. Methods We conducted a 12 month open-label pilot study to evaluate safety of oral DHA and its effects on serum alkaline phosphatase as a primary outcome measure in 23 patients with PSC. DHA was administered orally at 800 mg twice per day. Secondary outcomes included changes in other liver function tests and fibrosis biomarkers. Results A 1.7-fold increase in serum DHA levels was observed with supplementation. The mean alkaline phosphatase level (±S.E.) at baseline was 357.8 ± 37.1 IU compared to 297.1 ± 23.7 IU (P

Background Cystic fibrosis (CF) is characterized by an excessive inflammatory response in epithelial cells and macrophages. In CF mice, lung inflammation can be abrogated by oral treatment with docosahexaenoic acid (DHA). Since PPARs and LXRs are important regulators of inflammation and fatty acid metabolism in macrophages, we hypothesized that these pathways are dysregulated in CF macrophages and are corrected with DHA treatment. Methods Peritoneal macrophages were obtained from wild type and cftr −/− mice. LPS induced cytokine secretion and NFκB activity were analyzed with and without oral DHA treatment. The expression and activity of PPARα,γ, δ and LXRα were analyzed by RT-PCR and EMSA. Results LPS induced TNFα and IL-6 secretion and NFκB p65 activity were increased in CF macrophages. This was associated with low basal PPARγ expression and attenuated LPS induced induction of PPARδ, LXRα and ABCA1. DHA pretreatment in vivo decreased TNFα secretion and p65 activity, and increased PPARα and γ expression and function. The effects of DHA could be reproduced by PPAR agonists and blocked by a PPARα antagonist. Conclusion Impaired regulation of nuclear receptors may contribute to the abnormal LPS induced signaling in CF macrophages and is reversed by DHA.

The aim of the present study was to quantify absorption coefficients of specific fatty acids in preterm infants as a function of diet, formula or breast milk (BM), and postnatal age; to identify the fatty acid structural characteristics that determine optimal fatty acid absorption.Fatty acids from dietary and fecal samples were extracted and quantified by gas chromatography-mass spectroscopy. Fatty acid absorption coefficients (FA-CFAs) were calculated by comparing the total amount of fatty acids supplied by the diet to the amount quantified in the total fecal output during a 3-day period.A total of 18 infants (BM 8, formula 10) were studied at 2 weeks of age, and 20 infants (BM 10, formula 10) were studied at 6 weeks of age. FA-CFAs decreased with increasing carbon length in formula-fed infants at 2 and 6 weeks. Results were similar but less in magnitude in BM-fed infants at 2 weeks with no difference at 6 weeks.Preterm infants fed formula demonstrated lower FA-CFAs as a function of increasing carbon length. This is consistent with limited pancreatic lipase production and with lipase being present in BM but not in formula. The fact that this pattern was seen in BM-fed infants at 2 weeks but not 6 weeks of age suggests that intestinal immaturity may also play a role in impaired fatty acid absorption. These data highlight principles that need to be considered to optimize delivery and absorption of dietary long-chain polyunsaturated fatty acids in preterm infants.

Background: Feeding intolerance (FI), defined as delayed advancement to full enteral feedings, in the preterm infant leads to growth failure and may play a role in the pathogenesis of necrotizing enterocolitis, the leading gastrointestinal cause of death in the NICU. Methods: Serum gut hormone (GH) levels in 64 preterm infants born at < 30 weeks gestational age (GA) were measured on day of birth and at postnatal day 7 (Millipore, USA). Univariate linear regression was performed between GH levels and GA and between GH levels and days to full feedings. Results: Mean GA was 28.1 ± 1.3 wks and birthweight 1038.6 ± 276.6 gms. Leptin and gastrointestinal inhibitory peptide (GIP) on the day of birth (p=.04; p=.006) and leptin and pancreatic polypeptide (PP) at day 7 (p=.004; p=.01) were positively correlated with GA. Higher PP levels on the day of birth (p=.02) were correlated with FI while lower levels of GIP, polypeptide YY (PYY) and glucagon-like peptide 2 (GLP-2) at day 7 (p=.001; p=.0001; p=.03) were associ...

It is unknown why some patients with inflammatory bowel disease develop primary sclerosing cholangitis. We have recently shown that patients with primary sclerosing cholangitis have an increased prevalence of mutations in the gene responsible for cystic fibrosis (CFTR) compared with individuals with inflammatory bowel disease alone. Our aim was to examine whether induction of colitis by oral dextran leads to bile duct injury in mice heterozygous or homozygous for mutations in CFTR. The effect of oral administration of docosahexaenoic acid to correct a fatty acid imbalance associated with cystic fibrosis was also examined to determine whether this would prevent bile duct inflammation. Wild-type mice and mice heterozygous and homozygous for CFTR mutations were given dextran orally for 14 days to induce colitis. Bile duct injury was quantitated by blinded histological scoring and measurement of serum alkaline phosphatase activity. The effect of pretreatment with docosahexaenoic acid for 7 days was examined. Treatment of mice with 100 mg dextran/day for 9 days followed by 85 mg/day for 5 days resulted in a significant increase in bile duct injury as determined by histological scoring in homozygous cystic fibrosis mice compared with wild-type mice ( P = 0.005). The bile duct injury seen in cystic fibrosis mice was reflected in a threefold increase in serum alkaline phosphatase ( P = 0.0006). Pretreatment with oral docosahexaenoic acid decreased both histological evidence of bile duct injury and serum alkaline phosphatase levels. In the setting of colitis, loss of CFTR function leads to bile duct injury.

A long-standing impediment for applications of optical techniques in cellular biology is the inability to characterize subcellular structures whose dimensions are much less than about 1 /spl mu/m. In this paper, we describe a method based on light scattering spectroscopy that can find the size distribution of subcellular organelles as small as 100 nm with an accuracy of 20 nm. We report experiments using aqueous suspensions of subcellular organelles enriched in mitochondria, zymogen granules, and microsomes. From the observed light scattering spectra, we extract size distributions that are in excellent agreement with the results of electron microscopy. Further studies are underway to extract the shapes of organelles in addition to their sizes.

Background Morbidities of impaired immunity and dysregulated inflammation are common in preterm infants. Postnatal Intestinal development plays a critical role in the maturation of the immune system and is, in part, driven by exposure to an enteral diet. Objective The aim of this study was to evaluate the influence of the timing of the first enteral feeding on intestinal inflammation and risk of disease. Methods 130 infants

We have developed a novel optical method for observing submicron intracellular structures in living cells which is called confocal light absorption and scattering spectroscopic (CLASS) microscopy. It combines confocal microscopy, a well-established high-resolution microscopic technique, with light scattering spectroscopy (LSS). CLASS microscopy requires no exogenous labels and is capable of imaging and continuously monitoring individual viable cells, enabling the observation of cell and organelle functioning at scales on the order of 100 nm. In addition, it provides not only size information but also information about the biochemical and physical properties of the cell.

This article reports the development of an optical imaging technique, confocal light absorption and scattering spectroscopic (CLASS) microscopy, capable of noninvasively determining the dimensions and other physical properties of single subcellular organelles. CLASS microscopy combines the principles of light-scattering spectroscopy (LSS) with confocal microscopy. LSS is an optical technique that relates the spectroscopic properties of light elastically scattered by small particles to their size, refractive index, and shape. The multispectral nature of LSS enables it to measure internal cell structures much smaller than the diffraction limit without damaging the cell or requiring exogenous markers, which could affect cell function. Scanning the confocal volume across the sample creates an image. CLASS microscopy approaches the accuracy of electron microscopy but is nondestructive and does not require the contrast agents common to optical microscopy. It provides unique capabilities to study functions of viable cells, which are beyond the capabilities of other techniques.

We recently developed a new microscopic optical technique capable of noninvasive analysis of cell structure and cell dynamics on the submicron scale [1]. It combines confocal microscopy, a well-established high-resolution microscopic technique, with light scattering spectroscopy (LSS) and is called confocal light absorption and scattering spectroscopic (CLASS) microscopy. CLASS microscopy requires no exogenous labels and is capable of imaging and continuously monitoring individual viable cells, enabling the observation of cell and organelle functioning at scales on the order of 100 nm. To test the ability of CLASS microscopy to monitor cellular dynamics in vivo we performed experiments with human bronchial epithelial cells treated with DHA and undergoing apoptosis. The treated and untreated cells show not only clear differences in organelle spatial distribution but time sequencing experiments on a single cell show disappearance of certain types of organelles and change of the nuclear shape and density with the progression of apoptosis. In summary, CLASS microscopy provides an insight into metabolic processes within the cell and opens doors for the noninvasive real-time assessment of cellular dynamics. Noninvasive monitoring of cellular dynamics with CLASS microscopy can be used for a real-time dosimetry in a wide variety of medical and environmental applications that have no immediate observable outcome, such as photodynamic therapy, drug screening, and monitoring of toxins.

We recently developed a novel optical method for observing submicron intracellular structures in living cells which is called confocal light absorption and scattering spectroscopic (CLASS) microscopy. It combines confocal microscopy, a well-established high-resolution microscopic technique, with light scattering spectroscopy (LSS). CLASS microscopy requires no exogenous labels and is capable of imaging and continuously monitoring individual viable cells, enabling the observation of cell and organelle functioning at scales on the order of 100 nm with 10 nm accuracy. To demonstrate the ability of the CLASS microscope to monitor unstained living cells on submicrometer scale we studied human bronchial epithelial cells undergoing apoptosis. Fluorescence microscopy of living cells requires application of molecular markers which can affect normal cell functioning. CLASS microscopy is not affected by this avoiding potential interference of fluorescence molecular markers with cell processes. In addition, it provides not only size information but also information about the biochemical and physical properties of the cell. CLASS microscopy can provide unique capabilities for the study of cell interactions with the environment, cell reproduction and growth and other functions of viable cells, which are inaccessible by other techniques.

Primary sclerosing cholangitis (PSC) is associated with mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. As proof of concept that CFTR dysfunction plays a role in PSC, induction of colitis in cftr mice results in bile duct injury that can be prevented by pretreatment with docosahexaenoic acid (DHA).Determine whether 1) CFTR dysfunction in cftr mice through a reduction in peroxisome proliferator activated receptor (PPAR)alpha or gamma leads to bile duct injury and 2) whether DHA prevents bile duct injury through an increase in PPAR.Cftr and wild-type (WT) mice were treated with dextran sodium sulfate (DSS) to induce colitis with or without pretreatment with oral DHA. PPARalpha and gamma as well as tumor necrosis factor (TNF)alpha were analyzed in liver tissue. PPARalpha mice were also treated with DSS and histology examined.PPARgamma mRNA levels were low, with DSS suppressing mRNA levels equally in WT and cftr mice. PPARalpha levels were no different between cftr and WT litter mates by reverse-transcription polymerase chain reaction. After DSS, WT mice showed a 9.3-fold increase in PPARalpha mRNA levels and increased nuclear localization compared with no DSS (P0.05), with no increase seen in cftr mice. This was not caused by changes in TNFalpha. DHA treatment led to 7.0-fold increase in PPARalpha mRNA levels in cftr mice (P0.01). PPARalpha mice treated with DSS did not develop bile duct injury, indicating that PPARalpha alone is not sufficient to cause bile duct inflammation.DSS induced bile duct injury in cftr mice is associated with a defect in PPARalpha expression, which is reversed by DHA.

We report the development of a novel optical imaging technique, confocal light scattering spectroscopic microscopy, capable of non-invasively determining the dimensions and other physical properties of single subcellular organelles with 5 nm accuracy.

BACKGROUND Patients with cystic fibrosis have altered levels of plasma fatty acids. We previously demonstrated that arachidonic acid levels are increased and docosahexaenoic acid levels are decreased in affected tissues from cystic fibrosis-knockout mice. In this study we determined whether humans with mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene have a similar fatty acid defect in tissues expressing CFTR. METHODS Fatty acids from nasal- and rectal-biopsy specimens, nasal epithelial scrapings, and plasma were analyzed from 38 subjects with cystic fibrosis and compared with results in 13 obligate heterozygotes, 24 healthy controls, 11 subjects with inflammatory bowel disease, 9 subjects with upper respiratory tract infection, and 16 subjects with asthma. RESULTS The ratio of arachidonic to docosahexaenoic acid was increased in mucosal and submucosal nasal-biopsy specimens (P

To measure the changes in whole blood fatty acid levels in premature infants and evaluate associations between these changes and neonatal morbidities.This was a retrospective cohort study of 88 infants born at30 weeks' gestation. Serial fatty acid profiles during the first postnatal month and infant outcomes, including chronic lung disease (CLD), retinopathy of prematurity, and late-onset sepsis, were analyzed. Regression modeling was applied to determine the association between fatty acid levels and neonatal morbidities.Docosahexaenoic acid (DHA) and arachidonic acid levels declined rapidly in the first postnatal week, with a concomitant increase in linoleic acid levels. Decreased DHA level was associated with an increased risk of CLD (OR, 2.5; 95% CI, 1.3-5.0). Decreased arachidonic acid level was associated with an increased risk of late-onset sepsis (hazard ratio, 1.4; 95% CI, 1.1-1.7). The balance of fatty acids was also a predictor of CLD and late-onset sepsis. An increased linoleic acid:DHA ratio was associated with an increased risk of CLD (OR, 8.6; 95% CI, 1.4-53.1) and late-onset sepsis (hazard ratio, 4.6; 95% CI, 1.5-14.1).Altered postnatal fatty acid levels in premature infants are associated with an increased risk of CLD and late-onset sepsis.

We have developed a novel optical method for observing submicrometer intracellular structures in living cells, which is called confocal light absorption and scattering spectroscopic (CLASS) microscopy. It combines confocal microscopy, a well-established high-resolution microscopic technique, with light-scattering spectroscopy. CLASS microscopy requires no exogenous labels and is capable of imaging and continuously monitoring individual viable cells, enabling the observation of cell and organelle functioning at scales of the order of 100 nm.

Introduction Intragastru balloon systems have been used tot weight loss m obese patients. Because of a high complication rate this method was abandoned in the 1980s. Recently, a new generation of intragastric balloon system has been introduced in clinical use. We report the eNciency and tolerance of the Bioenterics lntragastric Balloon System (BIB). Methods Obese patients willing to reduce weight wetv oticred the possibility of weight reduction assistance by the BIB Initially a questionnaire was completed by all patients, inquiring about medical history, comorbidiv, dietary' habits and previous treatment t0r obesity. BIB placement was pertormed during upper endoscopy on an outpatient basis under intravenous sedation. The system was tilled with 500ml saline, coloured with methyienblue to identity, any leakage, A dieticians consultation was pertomled every month. lhe BIB was removed endoscopically aher 6 months Weight loss was a s se~d each month and alter removal ot the BIB. Results From July 2001 to November 2002, 60 Patients (19 male, 49 iemale, mean age 44.2 years (range 26 67)) underwent BIB iruplacement. Mean initial BMi was 38.6 kg/ru 2 (range 2952) witil a mean imtial excess weight of 41.8 kg (range 13-80) and nman initial %excess wright ot 3 5 8 (133-549) Iniual upper endoscopy was normal in 43 and abnormal in 17 patients (9 esophagitis, 3 small hiatal hernia, 4 gastritis, 3 duodenal ulcers, 2 patienl:s with double pattmlognes). In all 60 patients the insertion procedure was uneventful, in 2 patients the BIB had to be removed within the first week due to intolerance, in the retraining 58 patients the mean intmgastric residing time of tbe BIB was 6 6 months (range 4.5-9.0). There was no spomaneons loss ot the device. The mean weight loss was l l .2kg (range 035~ corresponding to a %excess weight loss of 30% (0-116.7). In the first week of treatment 48 patients (82%) suffered t~rom vomiting over a mean time of 3 7 days, heartburn occurred in 13 patients (22%) and hospitalisation was necessary m 2 patients Vomiting and heartburn requinng PPi-therapy lasted longer than one week in 8 (13%) and 14 (24%) patients, respectively No major complications like pertoration or obstruction were observed. Conclusion "[he insertion of BIB offers a highly efhcient method tot rapid weight reduction in obese patients Vomiting and retlux s}~nptoms are common within the tirst 4 days alter placement, but can be controlled by drug therapy Overall, the therapy is well tolerated, sate and associated with a low complication rate.