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1.  Favorable Changes in Cardiac Geometry and Function Following Gastric Bypass Surgery 
The objective of this study was to test the hypothesis that gastric bypass surgery (GBS) would favorably impact cardiac remodeling and function.
GBS is increasingly used to treat severe obesity, but there are limited outcome data.
We prospectively studied 423 severely obese patients undergoing GBS and a reference group of severely obese subjects that did not have surgery (n = 733).
At a 2-year follow up, GBS subjects had a large reduction in body mass index compared with the reference group (−15.4 ± 7.2 kg/m2 vs. −0.03 ± 4.0 kg/m2; p < 0.0001), as well as significant reductions in waist circumference, systolic blood pressure, heart rate, triglycerides, low-density lipoprotein cholesterol, and insulin resistance. High-density lipoprotein cholesterol increased. The GBS group had reductions in left ventricular (LV) mass index and right ventricular (RV) cavity area. Left atrial volume did not change in GBS but increased in reference subjects. In conjunction with reduced chamber sizes, GBS subjects also had increased LV midwall fractional shortening and RV fractional area change. In multivariable analysis, age, change in body mass index, severity of nocturnal hypoxemia, E/E', and sex were independently associated with LV mass index, whereas surgical status, change in waist circumference, and change in insulin resistance were not.
Marked weight loss in patients undergoing GBS was associated with reverse cardiac remodeling and improved LV and RV function. These data support the use of bariatric surgery to prevent cardiovascular complications in severe obesity.
PMCID: PMC3713780  PMID: 21292133
bariatric surgery; cardiac remodeling; diabetes; echocardiography; hypertension; left atrial volume; left ventricular hypertrophy; myocardial contraction; obesity
2.  Monitoring the progression of metastatic breast cancer on nanoporous silica chips 
Breast cancer accounted for 15 per cent of total cancer deaths in female patients in 2010. Although significant progress has been made in treating early-stage breast cancer patients, there is still no effective therapy targeting late-stage metastatic breast cancers except for the conventional chemotherapy interventions. Until effective therapy for later-stage cancers emerges, the identification of biomarkers for the early detection of tumour metastasis continues to hold the key to successful management of breast cancer therapy. Our study concentrated on the low molecular weight (LMW) region of the serum protein and the information it contains for identifying biomarkers that could reflect the ongoing physiological state of all tissues. Owing to technical difficulties in harvesting LMW species, studying these proteins/peptides has been challenging until now. In our study, we have recently developed nanoporous chip-based technologies to separate small proteins/peptides from the large proteins in serum. We used nanoporous silica chips, with a highly periodic nanostructure and uniform pore size distribution, to isolate LMW proteins and peptides from the serum of nude mice with MDA-MB-231 human breast cancer lung metastasis. By matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and biostatistical analysis, we were able to identify protein signatures unique to different stages of cancer development. The approach and results reported in this study possess a significant potential for the discovery of proteomic biomarkers that may significantly enhance personalized medicine targeted at metastatic breast cancer.
PMCID: PMC3318679  PMID: 22509065
nanoporous silica; breast cancer lung metastasis; low molecular weight protein; proteomic biomarker; early diagnosis
3.  Polymorphisms in the P2X7 receptor gene are associated with low lumbar spine bone mineral density and accelerated bone loss in post-menopausal women 
The P2X7 receptor gene (P2RX7) is highly polymorphic with five previously described loss-of-function (LOF) single-nucleotide polymorphisms (SNP; c.151+1G>T, c.946G>A, c.1096C>G, c.1513A>C and c.1729T>A) and one gain-of-function SNP (c.489C>T). The purpose of this study was to determine whether the functional P2RX7 SNPs are associated with lumbar spine (LS) bone mineral density (BMD), a key determinant of vertebral fracture risk, in post-menopausal women. We genotyped 506 post-menopausal women from the Aberdeen Prospective Osteoporosis Screening Study (APOSS) for the above SNPs. Lumbar spine BMD was measured at baseline and at 6–7 year follow-up. P2RX7 genotyping was performed by homogeneous mass extension. We found association of c.946A (p.Arg307Gln) with lower LS-BMD at baseline (P=0.004, β=−0.12) and follow-up (P=0.002, β=−0.13). Further analysis showed that a combined group of subjects who had LOF SNPs (n=48) had nearly ninefold greater annualised percent change in LS-BMD than subjects who were wild type at the six SNP positions (n=84; rate of loss=−0.94%/year and −0.11%/year, respectively, P=0.0005, unpaired t-test). This is the first report that describes association of the c.946A (p.Arg307Gln) LOF SNP with low LS-BMD, and that other LOF SNPs, which result in reduced or no function of the P2X7 receptor, may contribute to accelerated bone loss. Certain polymorphic variants of P2RX7 may identify women at greater risk of developing osteoporosis.
PMCID: PMC3330223  PMID: 22234152
P2RX7; LS-BMD; single-nucleotide polymorphisms
4.  Low Molecular Weight Protein Enrichment on Mesoporous Silica Thin Films for Biomarker Discovery 
The identification of circulating biomarkers holds great potential for non invasive approaches in early diagnosis and prognosis, as well as for the monitoring of therapeutic efficiency.1-3 The circulating low molecular weight proteome (LMWP) composed of small proteins shed from tissues and cells or peptide fragments derived from the proteolytic degradation of larger proteins, has been associated with the pathological condition in patients and likely reflects the state of disease.4,5 Despite these potential clinical applications, the use of Mass Spectrometry (MS) to profile the LMWP from biological fluids has proven to be very challenging due to the large dynamic range of protein and peptide concentrations in serum.6 Without sample pre-treatment, some of the more highly abundant proteins obscure the detection of low-abundance species in serum/plasma. Current proteomic-based approaches, such as two-dimensional polyacrylamide gel-electrophoresis (2D-PAGE) and shotgun proteomics methods are labor-intensive, low throughput and offer limited suitability for clinical applications.7-9 Therefore, a more effective strategy is needed to isolate LMWP from blood and allow the high throughput screening of clinical samples.
Here, we present a fast, efficient and reliable multi-fractionation system based on mesoporous silica chips to specifically target and enrich LMWP.10,11 Mesoporous silica (MPS) thin films with tunable features at the nanoscale were fabricated using the triblock copolymer template pathway. Using different polymer templates and polymer concentrations in the precursor solution, various pore size distributions, pore structures, connectivity and surface properties were determined and applied for selective recovery of low mass proteins. The selective parsing of the enriched peptides into different subclasses according to their physicochemical properties will enhance the efficiency of recovery and detection of low abundance species. In combination with mass spectrometry and statistic analysis, we demonstrated the correlation between the nanophase characteristics of the mesoporous silica thin films and the specificity and efficacy of low mass proteome harvesting. The results presented herein reveal the potential of the nanotechnology-based technology to provide a powerful alternative to conventional methods for LMWP harvesting from complex biological fluids. Because of the ability to tune the material properties, the capability for low-cost production, the simplicity and rapidity of sample collection, and the greatly reduced sample requirements for analysis, this novel nanotechnology will substantially impact the field of proteomic biomarker research and clinical proteomic assessment.
PMCID: PMC3466656  PMID: 22546927
Bioengineering;  Issue 62;  Nanoporous silica chip;  Low molecular weight proteomics;  Peptidomics;  MALDI-TOF mass spectrometry;  early diagnostics;  proteomics
5.  Receptors and effects of gut hormones in three osteoblastic cell lines 
BMC Physiology  2011;11:12.
In recent years the interest on the relationship of gut hormones to bone processes has increased and represents one of the most interesting aspects in skeletal research. The proportion of bone mass to soft tissue is a relationship that seems to be controlled by delicate and subtle regulations that imply "cross-talks" between the nutrient intake and tissues like fat. Thus, recognition of the mechanisms that integrate a gastrointestinal-fat-bone axis and its application to several aspects of human health is vital for improving treatments related to bone diseases. This work analysed the effects of gut hormones in cell cultures of three osteoblastic cell lines which represent different stages in osteoblastic development. Also, this is the first time that there is a report on the direct effects of glucagon-like peptide 2, and obestatin on osteoblast-like cells.
mRNA expression levels of five gut hormone receptors (glucose-dependent insulinotropic peptide [GIP], glucagon-like peptide 1 [GLP-1], glucagon-like peptide 2 [GLP-2], ghrelin [GHR] and obestatin [OB]) were analysed in three osteoblastic cell lines (Saos-2, TE-85 and MG-63) showing different stages of osteoblast development using reverse transcription and real time polymerase chain reaction. The responses to the gut peptides were studied using assays for cell viability, and biochemical bone markers: alkaline phosphatase (ALP), procollagen type 1 amino-terminal propeptides (P1NP), and osteocalcin production.
The gut hormone receptor mRNA displayed the highest levels for GIP in Saos-2 and the lowest levels in MG-63, whereas GHR and GPR39 (the putative obestatin receptor) expression was higher in TE-85 and MG-63 and lower in Saos-2. GLP-1 and GLP-2 were expressed only in MG-63 and TE-85. Treatment of gut hormones to cell lines showed differential responses: higher levels in cell viability in Saos-2 after GIP, in TE-85 and MG-63 after GLP-1, GLP-2, ghrelin and obestatin. ALP showed higher levels in Saos-2 after GIP, GHR and OB and in TE-85 after GHR. P1NP showed higher levels after GIP and OB in Saos-2. Decreased levels of P1NP were observed in TE-85 and MG-63 after GLP-1, GLP-2 and OB. MG-63 showed opposite responses in osteocalcin levels after GLP-2.
These results suggest that osteoblast activity modulation varies according to different development stage under different nutrition related-peptides.
PMCID: PMC3162581  PMID: 21801348
6.  Phosphorylated Dihydroceramides from Common Human Bacteria Are Recovered in Human Tissues 
PLoS ONE  2011;6(2):e16771.
Novel phosphorylated dihydroceramide (PDHC) lipids produced by the periodontal pathogen Porphyromonas gingivalis include phosphoethanolamine (PE DHC) and phosphoglycerol dihydroceramides (PG DHC) lipids. These PDHC lipids mediate cellular effects through Toll-like receptor 2 (TLR2) including promotion of IL-6 secretion from dendritic cells and inhibition of osteoblast differentiation and function in vitro and in vivo. The PE DHC lipids also enhance (TLR2)-dependent murine experimental autoimmune encephalomyelitis (EAE), a model for multiple sclerosis. The unique non-mammalian structures of these lipids allows for their specific quantification in bacteria and human tissues using multiple reaction monitoring (MRM)-mass spectrometry (MS). Synthesis of these lipids by other common human bacteria and the presence of these lipids in human tissues have not yet been determined. We now report that synthesis of these lipids can be attributed to a small number of intestinal and oral organisms within the Bacteroides, Parabacteroides, Prevotella, Tannerella and Porphyromonas genera. Additionally, the PDHCs are not only present in gingival tissues, but are also present in human blood, vasculature tissues and brain. Finally, the distribution of these TLR2-activating lipids in human tissues varies with both the tissue site and disease status of the tissue suggesting a role for PDHCs in human disease.
PMCID: PMC3037954  PMID: 21347306
7.  The effects of P2X7 receptor antagonists on the formation and function of human osteoclasts in vitro 
Purinergic Signalling  2010;6(3):307-315.
The P2X7 receptor (P2X7R) has been implicated in the process of multinucleation and cell fusion. We have previously demonstrated that blockade of P2X7Rs on osteoclast precursors using a blocking antibody inhibited multinucleated osteoclast formation in vitro, but that P2X7R KO mice maintain the ability to form multinucleated osteoclasts. This apparent contradiction of the role the P2X7R plays in multinucleation has prompted us to examine the effect of the most commonly used and recently available P2X7R antagonists on osteoclast formation and function. When added to recombinant RANKL and M-CSF human blood monocytes cultures, all but one compound, decreased the formation and function of multinucleated TRAP-positive osteoclasts in a concentration-dependent manner. These data provide further evidence for the role of the P2X7R in the formation of functional human multinucleated osteoclasts and highlight the importance of selection of antagonists for use in long-term experiments.
PMCID: PMC2947658  PMID: 21103214
P2X7; ATP; Osteoclast; P2 receptor; Resorption; Formation; Fusion
8.  Reduced Eukaryotic Initiation Factor 2B ε-Subunit Expression Suppresses the Transformed Phenotype of Cells Overexpressing the Protein 
Cancer research  2008;68(21):8752-8760.
Eukaryotic initiation factor 2B (eIF2B), a five subunit guanine nucleotide exchange factor (GEF), plays a key role in the regulation of mRNA translation. Expression of its ε-subunit is specifically upregulated in certain conditions associated with increased cell growth. Therefore, the purpose of the present study was to examine the effect of repressing eIF2Bε expression on growth rate, protein synthesis, and other characteristics of two tumorigenic cell lines that display upregulated expression of the ε-subunit. Experiments were designed to compare spontaneously transformed fibroblasts (TMEF’s) to TMEFs infected with a lentivirus containing a short hairpin (sh)RNA directed against eIF2Bε. Cells expressing the shRNA displayed a reduction in eIF2Bε abundance to 30% of the value observed in uninfected TMEF’s with no change in the expression of any of the other four subunits. The repression of eIF2Bε expression was accompanied by reductions in GEF activity and global rates of protein synthesis. Moreover, repressed eIF2Bε expression led to marked reductions in cell growth rate in culture, colony formation in soft agar, and tumor progression in nude mice. Similar results were obtained in MCF-7 human breast cancer cells in which eIF2Bε expression was repressed through transient transfection with a siRNA directed against the ε-subunit. Overall, the results support a role for eIF2Bε in the regulation of cell growth and suggest that it might represent a therapeutic target for the treatment of human cancer.
PMCID: PMC2664734  PMID: 18974117
eIF2B5; transformation; shRNA; siRNA
9.  Activation of the Mammalian Target of Rapamycin Complex 1 is Both Necessary and Sufficient to Stimulate Eukaryotic Initiation Factor 2Bε mRNA Translation and Protein Synthesis 
In a previous study we demonstrated a requirement for activation of mTORC1 in the stimulation of eIF2Bε mRNA translation in skeletal muscle in response to resistance exercise. Although that study established the necessity of mTORC1 activation, the experimental model used did not lend itself readily to address the question of whether or not mTORC1 activation was sufficient to produce the response. Therefore, the present study was designed to address the sufficiency of mTORC1 activation, using cultures of Rat2 fibroblasts in which mTORC1 signaling was repressed by serum/leucine-depletion and stimulated by repletion of leucine and/or IGF-1. Repletion with leucine and IGF-1 caused a shift of eIF2Bε mRNA into actively translating polysomes and a stimulation of new eIF2Bε protein synthesis, but had no effect on mRNAs encoding the other four eIF2B subunits. Stimulation of eIF2Bε translation was reversed by pre-treatment with the mTORC1 inhibitor rapamycin. Exogenous overexpression of FLAG-Rheb, a proximal activator of mTORC1, also caused a re-distribution of eIF2Bε mRNA into polysomes and a stimulation of eIF2Bε protein synthesis. The stimulation of eIF2Bε mRNA translation occurred in the absence of any effect on eIF2Bε mRNA abundance. RNAi-mediated knockdown of eIF2Bε resulted in reduced cellular proliferation, a result that phenocopied the known cytostatic effect of mTORC1 repression. Overall the results demonstrate that activation of mTORC1 is both necessary and sufficient to stimulate eIF2Bε mRNA translation and that this response may represent a novel mechanism through which mTORC1 can affect mRNA translation initiation, rates of protein synthesis, and cellular growth/proliferation.
PMCID: PMC2601645  PMID: 18556237
mTOR; eIF2B; Rheb; mRNA translation
10.  ABC of eyes: Injury to the eye 
BMJ : British Medical Journal  2004;328(7440):644.
PMCID: PMC381182  PMID: 15016709
12.  Adenovirus Susceptibility to Human Interferon During One-Step Replication 1 
Infection and Immunity  1972;5(6):905-908.
Susceptibility of adenovirus types 2, 7, and 12 to human interferon was measured in three human diploid cell strains during a single-cycle infection. Although the relative susceptibility of adenovirus to interferon varied in these cell strains, the final yield of each type in each cell strain decreased as the interferon dose increased. On the other hand, wide difference in interferon susceptibility of adenoviruses and vesicular stomatitis virus (VSV) was noted, as interferon doses above 100 units profoundly inhibited VSV but not the adenoviruses.
PMCID: PMC422461  PMID: 4344095

Results 1-12 (12)