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1.  Effects of Short Read Quality and Quantity on a de novo Vertebrate Transcriptome Assembly✰ 
For many researchers, next generation sequencing data holds the key to answering a category of questions previously unassailable. One of the important and challenging steps in achieving these goals is accurately assembling the massive quantity of short sequencing reads into full nucleic acid sequences. For research groups working with non-model or wild systems, short read assembly can pose a significant challenge due to the lack of pre-existing EST or genome reference libraries. While many publications describe the overall process of sequencing and assembly, few address the topic of how many and what types of reads are best for assembly. The goal of this project was use real world data to explore the effects of read quantity and short read quality scores on the resulting de novo assemblies. Using several samples of short reads of various sizes and qualities we produced many assemblies in an automated manner. We observe how the properties of read length, read quality, and read quantity affect the resulting assemblies and provide some general recommendations based on our real-world data set.
PMCID: PMC3223268  PMID: 21651990
NGS; short read; quality; Phred; assembly; quantity; Velvet
2.  Autophagy promotes T-cell survival through degradation of proteins of the cell death machinery 
Cell Death and Differentiation  2011;19(1):144-152.
Autophagy is implicated in regulating cell death in activated T cells, but the underlying mechanism is unclear. Here, we show that inhibition of autophagy via Beclin 1 gene deletion in T cells leads to rampant apoptosis in these cells upon TCR stimulation. Beclin 1-deficient mice fail to mount autoreactive T-cell responses and are resistant to experimental autoimmune encephalomyelitis. Compared with Th17 cells, Th1 cells are much more susceptible to cell death upon Beclin 1 deletion. Cell death proteins are highly increased in Beclin 1-deficient T cells and inhibition of caspases and genetic deletion of Bim reverse apoptosis. In addition, p62/sequestosome 1 binds to caspase-8 but does not control levels of procaspase-8 or other cell death-related proteins. These results establish a direct role of autophagy in inhibiting the programmed cell death through degradation of apoptosis proteins in activated T cells.
PMCID: PMC3252822  PMID: 21660048
T cell; Th1 cell; apoptosis; autophagy; caspase; Beclin 1
3.  Castleman's Disease: A Case Report of the Unicentric Type 
Case Reports in Surgery  2012;2012:175272.
Castleman's disease, or angiofollicular lymph node hyperplasia, is a relatively rare disorder characterized by the benign proliferation of lymphoid tissue related to the chronic human herpes virus 8 (HHV-8) infection and the human immunodeficiency virus (HIV). Two clinical entities have been described: a unicentric presentation with the disease confined to a single anatomic lymph node and a multicentric presentation characterized by generalized lymphadenopathy and a more aggressive clinical course. Also, three histopathological subtypes have been described: hyaline-vascular, plasma cell, and a mixed variant. Preoperative diagnosis of hyaline-vascular Castleman's disease is difficult, and the definitive result is based on postoperative pathological findings. The gold standard therapy is the complete surgical excision.
PMCID: PMC3443986  PMID: 22991681
4.  Global Oral Health Inequalities 
Advances in Dental Research  2011;23(2):207-210.
Despite impressive worldwide improvements in oral health, inequalities in oral health status among and within countries remain a daunting public health challenge. Oral health inequalities arise from a complex web of health determinants, including social, behavioral, economic, genetic, environmental, and health system factors. Eliminating these inequalities cannot be accomplished in isolation of oral health from overall health, or without recognizing that oral health is influenced at multiple individual, family, community, and health systems levels. For several reasons, this is an opportune time for global efforts targeted at reducing oral health inequalities. Global health is increasingly viewed not just as a humanitarian obligation, but also as a vehicle for health diplomacy and part of the broader mission to reduce poverty, build stronger economies, and strengthen global security. Despite the global economic recession, there are trends that portend well for support of global health efforts: increased globalization of research and development, growing investment from private philanthropy, an absolute growth of spending in research and innovation, and an enhanced interest in global health among young people. More systematic and far-reaching efforts will be required to address oral health inequalities through the engagement of oral health funders and sponsors of research, with partners from multiple public and private sectors. The oral health community must be “at the table” with other health disciplines and create opportunities for eliminating inequalities through collaborations that can harness both the intellectual and financial resources of multiple sectors and institutions.
PMCID: PMC3144034  PMID: 21490232
global; oral health; inequalities; disparities; research; funding
7.  An anthropological approach to the evaluation of preschool children exposed to pesticides in Mexico. 
Environmental Health Perspectives  1998;106(6):347-353.
In this comparative study, we compensated for many of the known variables that influence children's growth and development by selecting two groups of 4-5-year-old Yaqui children who reside in the Yaqui Valley of northwestern Mexico. These children share similar genetic backgrounds, diets, water mineral contents, cultural patterns, and social behaviors. The major difference was their exposure to pesticides. Pesticides have been applied to the agricultural area of the valley since the late 1940s. In 1990, high levels of multiple pesticides were found in the cord blood of newborns and in breast milk. Building on anthropological methods for rapid rural appraisal of problems within the environment, a Rapid Assessment Tool for Preschool Children (RATPC) was developed to measure growth and development. The children of the agrarian region were compared to children living in the foothills, where pesticide use is avoided. The RATPC measured varied aspects of physical growth and abilities to perform, or function in, normal childhood activities. No differences were found in growth patterns. Functionally, the exposed children demonstrated decreases in stamina, gross and fine eye-hand coordination, 30-minute memory, and the ability to draw a person. The RATPC also pointed out areas in which more in-depth research on the toxicology of pesticides would be valuable.
PMCID: PMC1533004  PMID: 9618351
8.  Transgenic mice expressing soluble tumor necrosis factor-receptor are protected against bone loss caused by estrogen deficiency. 
Journal of Clinical Investigation  1997;99(7):1699-1703.
To evaluate the role of tumor necrosis factor (TNF alpha) in bone loss resulting from estrogen deficiency, the effects of ovariectomy were explored in six-month-old transgenic mice expressing high blood levels of a soluble TNF receptor type I (sTNFR1)-FcIgG3 fusion protein, which neutralizes TNF alpha, and in their nontransgenic littermates used as controls. These transgenic mice were identical to control mice in bone mass (evaluated by bone mineral density and content) and strength. 12 weeks after ovariectomy, the decrease in bone mass and increase in osteocalcin (marker of bone turnover) found in control mice were not observed in transgenic mice, which were not different from sham-operated mice, transgenic or not. This observation suggests a critical role for TNF alpha in the pathogenesis of bone loss induced by estrogen deficiency, a common cause of morbidity in postmenopausal women.
PMCID: PMC507990  PMID: 9120014
9.  Evidence for a protective role of tumor necrosis factor in the acute phase of Trypanosoma cruzi infection in mice. 
Infection and Immunity  1997;65(2):457-465.
A possible role for tumor necrosis factor (TNF) alpha during Trypanosoma cruzi infection was explored by using transgenic mice expressing in blood high levels of a soluble TNFR1-FcIgG3 fusion protein, which neutralizes the effects of TNF in vivo. Nontransgenic littermates were used as controls. The transgenic mice showed high susceptibility to T. cruzi infection. Inocula sublethal for control mice resulted in over 80% mortality associated with higher levels of parasites in the blood. In histological sections of the hearts of transgenic mice, large parasitic clusters without inflammatory cell infiltrates around the parasites were seen, while smaller parasitic clusters associated with leukocytes were seen in control mice. No difference in specific antibody response or lymphocyte composition of the spleen was found between transgenic and control mice, although the unresponsiveness of spleen cells to concanavalin A stimulation in vitro, typical of the acute phase of T. cruzi infection, was less pronounced in transgenic mice. Infected transgenic mice produced higher levels of gamma interferon than did control mice. These results confirm that TNF is involved in mechanisms leading to parasite clearance and protection from death in the acute phase of T. cruzi infection. More importantly, the data reveal that TNF is necessary for the establishment of effective tissue inflammation and parasite load control in acute experimental Chagas' disease myocarditis.
PMCID: PMC174617  PMID: 9009297
10.  Inhibition of submandibular and lacrimal gland infiltration in nonobese diabetic mice by transgenic expression of soluble TNF-receptor p55. 
Journal of Clinical Investigation  1996;98(4):954-961.
Besides a prominent mononuclear cell infiltration of the islets of Langerhans, nonobese diabetic (NOD) mice also show massive cellular infiltrates of the submandibular and lacrimal glands concomitant with histological signs of tissue damage. To obtain insights into the mechanisms operative during the initiation and progression of tissue damage, we followed by in situ hybridization the appearance of cells containing mRNA of the gene encoding the proinflammatory cytokine TNF-alpha in the cellular infiltrates. Cells expressing TNF-alpha are mainly located in infiltrates, are absent in nonaffected glands, and are preferentially found among CD4 T cells. Secretion of TNF-alpha by gland-infiltrating cells was confirmed by an ELISPOT procedure. Direct evidence for an instrumental role of TNF-alpha in initiation and progression of submandibular and lacrimal gland infiltration is provided by the observed significant reduction in the extent of infiltration in nonobese diabetic mice transgenic for a soluble TNF receptor p55 fused to the Fc part of human IgG3. This protection from infiltration is paralleled by decreased expression of the adhesion molecules ICAM-1 and VCAM-1 in submandibular and lacrimal glands. These data suggest a central role of TNF-alpha in the initiation and progression of autoimmune tissue destruction of salivary glands and indicate beneficial effects of soluble TNF receptors in the treatment of organ-specific autoimmune diseases.
PMCID: PMC507510  PMID: 8770867
11.  Expression of a tumor necrosis factor-alpha transgene in murine lung causes lymphocytic and fibrosing alveolitis. A mouse model of progressive pulmonary fibrosis. 
Journal of Clinical Investigation  1995;96(1):250-259.
The murine TNF-alpha gene was expressed under the control of the human surfactant protein SP-C promoter in transgenic mice. A number of the SP-C TNF-alpha mice died at birth or after a few weeks with very severe lung lesions. Surviving mice transmitted a pulmonary disease to their offspring, the severity and evolution of which was related to the level of TNF-alpha mRNA in the lung; TNF-alpha RNA was detected in alveolar epithelium, presumably in type II epithelial cells. In a longitudinal study of two independent mouse lines, pulmonary pathology, at 1-2 mo of age, consisted of a leukocytic alveolitis with a predominance of T lymphocytes. Leukocyte infiltration was associated with endothelial changes and increased levels of mRNA for the endothelial adhesion molecule VCAM-1. In the following months, alveolar spaces enlarged in association with thickening of the alveolar walls due to an accumulation of desmin-containing fibroblasts, collagen fibers, and lymphocytes. Alveolar surfaces were lined by regenerating type II epithelial cells, and alveolar spaces contained desquamating epithelial cells in places. Platelet trapping in the damaged alveolar capillaries was observed. Pulmonary pathology in the SP-C TNF-alpha mice bears a striking resemblance to human idiopathic pulmonary fibrosis, in which increased expression of TNF-alpha in type II epithelial cells has also been noted. These mice provide a valuable animal model for understanding the pathogenesis of pulmonary fibrosis and exploring possible therapeutic approaches.
PMCID: PMC185196  PMID: 7542280
12.  Ganciclovir prophylaxis decreases frequency and severity of cytomegalovirus disease in seropositive liver transplant recipients treated with OKT3 monoclonal antibodies. 
Antimicrobial Agents and Chemotherapy  1993;37(11):2490-2492.
The efficacy of ganciclovir, given prophylactically, to prevent cytomegalovirus-related disease was evaluated in liver transplant recipients, mostly seropositive, under treatment with OKT3 monoclonal antibodies. The incidence of cytomegalovirus disease and visceral involvement was reduced, respectively, from 52 and 36% in the control group to 12 and 8% in the ganciclovir-treated patients. Leukopenia was a frequent (32%) side effect of ganciclovir administration.
PMCID: PMC192416  PMID: 8285641
13.  Uptake and intracellular activity of an optically active ofloxacin isomer in human neutrophils and tissue culture cells. 
The penetration of an optically active ofloxacin isomer [(-)-ofloxacin] into human neutrophils and different tissue culture cells (HEp-2, McCoy, MDCK, and Vero) was studied and compared with that of ofloxacin by a fluorometric assay. The cellular-to-extracellular-concentration ratios (C/E) of (-)-ofloxacin were always higher than 6, significantly greater than those of ofloxacin at extracellular concentrations of 5 and 10 mg/liter. The penetration of (-)-ofloxacin and ofloxacin was doubled when neutrophils were stimulated by phorbol myristate acetate but not affected after ingestion of opsonized Staphylococcus aureus. The C/E ratios of (-)-ofloxacin and ofloxacin for different tissue culture epithelial cells and fibroblasts were lower than those of neutrophils but still higher than 2. Both compounds produced a significant reduction in viable intraphagocytic S. aureus during 3 h of exposure to antimicrobial agents. We conclude that (-)-ofloxacin appears to reach higher intracellular concentrations than ofloxacin, remaining active inside the neutrophils.
PMCID: PMC171573  PMID: 2327777
14.  Fluorometric measurement of ofloxacin uptake by human polymorphonuclear leukocytes. 
A fluorometric assay, based on the natural fluorescence of the quinolone nucleus, was used to determine the uptake of ofloxacin by human polymorphonuclear leukocytes. The ratio of cellular concentration to extracellular concentration (C/E) at 20 min and 37 degrees C was 7.2, using an extracellular concentration of 5 micrograms/ml. Uptake was rapid and was not affected by pH (5 to 9), but required elevated environmental temperature and cell viability. The metabolic inhibitors sodium fluoride and sodium cyanide significantly decreased the uptake of ofloxacin. The penetration of ofloxacin was not affected by the presence of glucose or adenosine, but was decreased by L-amino acids (lysine, leucine, and glycine). These results suggest that ofloxacin could be transported via an amino acid transport system and that the fluorometric assay is a useful method for determining the intracellular penetration of fluoroquinolones, avoiding the use of radiolabeled antimicrobial agents.
PMCID: PMC172508  PMID: 2751280
15.  Establishment of two rabbit mammary epithelial cell lines with distinct oncogenic potential and differentiated phenotype after microinjection of transforming genes. 
Molecular and Cellular Biology  1986;6(6):1974-1982.
The goal of this work was to establish an assay for transformation of epithelial cells. Two epithelial cell lines were obtained after microinjecting transforming genes into primary rabbit mammary secretory cells. The cell lines were analyzed for their oncogenic potential and for the maintenance of a differentiated phenotype. A fully transformed cell line, which retained epithelial cell organization, was obtained by coinjecting simian virus 40 DNA and the activated human c-Ha-ras gene. The proliferation rate of these cells was high, with a doubling time of 16 h. Their growth was anchorage independent, and they had lost contact inhibition. The cells were tumorigenic in nude mice, but had no metastatic potential. Both microinjected DNAs were efficiently transcribed and translated, in contrast to the casein genes, which were expressed in primary cells but not in the transformed cell line. An immortalized cell line established after injection with simian virus 40 DNA alone was characterized by a moderate rate of proliferation with a doubling time of approximately 30 h. The growth of these cells was contact inhibited and anchorage dependent. The cells were not tumorigenic in nude mice. The viral DNA was expressed during early passages, as shown by the presence of the large T antigen in cell nuclei, but not at later passages. A high number of lactogenic hormone receptors were found associated with the cell surface. Despite the presence of these receptors, no induction of genes coding for milk proteins was observed after addition of prolactin. These data demonstrate that this assay system can be used to assess the immortalizing and transforming potential of candidate oncogenes in epithelial cells.
PMCID: PMC367736  PMID: 3023914
16.  In vitro activity of WIN 49375 compared with those of other antibiotics in isolates from cancer patients. 
The activity of WIN 49375 [6-fluoro-1, 4-dihydro-1-(methylamino)-7-(4-methyl-1-piperazinyl)-4-oxo-3- quinolinecarboxylic acid], a new synthetic quinolone, was tested in vitro against 587 clinical isolates. The MICs for 90% of isolates of Escherichia coli, Klebsiella pneumoniae, and Enterobacter cloacae were 0.20, 1.56, and 0.39 microgram/ml, respectively. The MICs for 90% of isolates of Pseudomonas aeruginosa and Serratia marcescens were both 3.12 micrograms/ml. WIN 49375 was minimally active against gram-positive cocci. Its in vitro activity suggests that it may be useful for the treatment of gram-negative bacillary infections.
PMCID: PMC176185  PMID: 6508271
17.  In vitro activities of new beta-lactam antibiotics against Acinetobacter spp. 
The in vitro activities of new beta-lactam antibiotics was studied and compared with those of other known agents against 51 clinical isolates of Acinetobacter calcoaceticus subsp. anitratus and 23 isolates of A. calcoaceticus subsp. lwoffi. Of the new beta-lactam antibiotics, imipemide (N-formimidoyl thienamycin), ceftazidime, ceftizoxime, ceftriaxone, and piperacillin demonstrated good activities. The minimal inhibitory concentrations for A. calcoaceticus subsp. lwoffi were lower than those obtained for A. calcoaceticus subsp. anitratus.
PMCID: PMC185157  PMID: 6638992
18.  Multiple-dose pharmacokinetics of ceftazidime in cancer patients. 
The pharmacokinetic parameters of ceftazidime were determined in 25 patients with neoplastic diseases. A group of 13 patients each received 1 g of ceftazidime over 15 min as a single intravenous dose. The mean peak drug concentration in serum was 77.4 micrograms/ml, and the serum half-life was 144 min. Urinary excretion at 12 h was 64.5%. These 13 patients also received 2 g of ceftazidime over 15 min at 8-h intervals for 7 or 8 days. The mean peak drug concentrations in serum were 103.6 micrograms/ml on day 1 and 87 micrograms/ml on day 7 or 8. A second group of 12 patients received 1 g of ceftazidime over 30 min, followed by 1 g over 2 h and every 4 h thereafter. The mean peak drug concentration in serum at 30 min was 57.5 micrograms/ml. Average peak and trough levels obtained on day 2 or 3 were 65 and 34.8 micrograms/ml, respectively, and remained in this range thereafter. The above-mentioned dose schedules produced high and adequate ceftazidime concentrations in serum.
PMCID: PMC185126  PMID: 6357067
19.  Evaluation of Urinary Elimination of N-Acetyl-β-Glucosaminidase in Healthy Volunteers Treated with Dibekacin or Gentamicin 
The urinary excretion of N-acetyl-β-glucosaminidase was studied in healthy subjects during and after treatment with aminoglycosides. In terms of this parameter dibekacin appeared to be less nephrotoxic than gentamicin.
PMCID: PMC182065  PMID: 7114833

Results 1-19 (19)