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Initial Experiments (initial + experiment)
Selected AbstractsOntogeny of energy homeostatic pathways via neuroendocrine signaling in Atlantic salmonDEVELOPMENTAL NEUROBIOLOGY, Issue 9 2010Anne-Grethe Gamst Moen Abstract Leptin and ghrelin are known to regulate energy homeostasis via hypothalamic neuropeptide signaling in mammals. Recent studies have discovered that these hormones exist in teleosts, however, very little is known concerning their role during teleost ontogeny. Here, we have examined the steady state levels of leptins, ghrelins, their target neuropetides and several growth factors during Atlantic salmon development. Initial experiments revealed differential expression of leptin genes and ghrelin isoforms during embryogenesis. In larvae, equal upregulation of ghrl1 and ghrl2 was observed just prior to exogenous feeding while a surge of lepa1 occurred one week after first-feeding. Subsequent dissection of the embryos and larvae showed that lepa1, cart, pomca1, and agrp are supplied as maternal transcripts. The earliest zygotic expression was observed for lepa1 and cart at 320 day degrees. By 400 day degrees, this expression was localized to the head and coincided with upregulation of ghrl2 and npy. Over the hatching period growth factor signaling predominated. The ghrelin surge prior to first-feeding was exclusively localized in the internal organs and coincided with upregulation of npy and agrp in the head and agrp in the trunk. One week after exogenous feeding was established major peaks were detected in the head for lepa1 and pomca1 with increasing levels of cart, while lepa1 was also significantly expressed in the trunk. By integrating theses data into an ontogenetic model, we suggest that the mediation of Atlantic salmon energy homeostatic pathways via endocrine and neuropeptide signaling retains putative features of the mammalian system. © 2010 Wiley Periodicals, Inc. Develop Neurobiol 70: 649,658, 2010 [source] Inverse relationship between seizure expression and extrasynaptic NMDAR function following chronic NMDAR inhibitionEPILEPSIA, Issue 2010Suzanne B. Bausch Summary We showed previously that electrographic seizures involving dentate granule cells in organotypic hippocampal slice cultures were dramatically reduced following chronic treatment with the NR2B-selective antagonist, Ro25,6981, but were increased following chronic treatment with the high-affinity competitive antagonist, D(-)-2-amino-5-phosphonopentanoic acid (D-APV). To begin to investigate the potential mechanisms underlying the differential effects of N -methyl- d -aspartate receptor (NMDAR) antagonists on seizures, electrophysiologic experiments were conducted in dentate granule cells in hippocampal slice cultures treated for the entire 17,21 day culture period with vehicle, Ro25,6981 or D-APV. Initial experiments revealed a lack of an association between miniature excitatory postsynaptic current (mEPSC) measures and seizures suggesting that shifts in mEPSC were unlikely to account for the differential effects of D-APV and Ro25,6981 on seizures. However, the amplitude of tonic NMDAR-mediated currents was reduced in cultures treated chronically with D-APV and dramatically enhanced in cultures treated chronically with Ro25,6981. Because tonic NMDAR currents are mediated primarily by extrasynaptic NMDAR, these data show an inverse relationship between changes in extrasynaptic NMDAR function and alterations in seizure expression. [source] In vivo post-transcriptional regulation of CD154 in mouse CD4+ T cellsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 8 2009Stefano Vavassori Abstract Interactions between CD40 and its ligand CD154 are involved in the progression of both cell mediated and innate immunity. These interactions are brought about by the transient expression of CD154 on activated CD4+ T cells, which is regulated, in part, at the level of mRNA turnover. Here we have focused on analyzing the pattern of post-transcriptional regulation in mouse CD4+ T cells in response to activation. Initial experiments identify a region of the murine CD154 mRNA that binds a polypyrimidine tract-binding protein-containing complex (mComplex I), which is activation-dependent and binds to a single CU-rich site within the 3, uTR Subsequent findings demonstrate that in vivo polyclonal activation of T cells leads to a pattern of differential CD154 mRNA stability that is directly dependent on extent of activation. Furthermore, in vitro activation of antigen-primed T cells shows that the CD154 mRNA half-life increases relative to that of unprimed cells. Importantly, this is the first report demonstrating that the regulation of CD154 in vivo is connected to an activation-induced program of mRNA decay and thus provides strong evidence for post-transcriptional mechanisms having a physiological role in regulating CD154 expression during an ongoing immune response. [source] Systematic determination of ion score cutoffs based on calculated false positive rates: application for identifying ubiquitinated proteins by tandem mass spectrometryJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 3 2008Julian Vasilescu Abstract We report a simple approach for determining ion score cutoffs that permit the confident identification of ubiquitinated proteins by tandem mass spectrometry (MS/MS). Initial experiments involving the analysis of gel bands containing multi-Ubiquitin chains with quadrupole time-of-flight and quadrupole ion trap mass spectrometers revealed that standard ion score cutoffs used for database searching were not sufficiently stringent. We also found that false positive and false negative rates (FPR and FNR) varied significantly depending on the cutoff scores used and that appropriate cutoffs could only be determined following a systematic evaluation of false positive rates. When standard cutoff scores were used for the analysis of complex mixtures of ubiquitinated proteins, unacceptably high FPR were observed. Finally, we found that FPR for ubiquitinated proteins are affected by the size of the protein database that is searched. These observations may be applicable for the study of other post-translational modifications. Copyright © 2007 John Wiley & Sons, Ltd. [source] Analytical investigation of styrax and benzoin balsams by HPLC- PAD-fluorimetry and GC-MSPHYTOCHEMICAL ANALYSIS, Issue 4 2008Michael Hovaneissian Abstract An HPLC and GC study has been conducted on the aromatic oleoresins styrax and benzoin produced by several American, Mediterranean and East-Asian trees, and widely used in ancient civilisations for their therapeutic and odoriferous properties. Initial experiments were performed by HPLC-PAD-fluorimetry for the analysis of several aromatic components, and then completed by GC-MS for the characterisation of both aromatic and triterpenic derivatives. In this work, it was crucial to isolate from fresh natural exudates, and to characterise by two-dimensional NMR, some of the major constituents in order to extend the standard molecular pool prior to chromatographic identifications. This study reveals coniferyl benzoate as an excellent distinctive fluorescent biomarker of Siam benzoin substrate. It also confirms that fluorimetric-coupled detection is a powerful analytical tool for the identification of compounds in Hamamelidaceae extracts that are almost undetectable by UV. GC-MS was successfully applied to the determination of the botanical origin of Sumatra benzoin, and to the identification of lupeol [3, -lup-20(29)-en-3-ol] for the first time in such balsam-type materials. Copyright © 2007 John Wiley & Sons, Ltd. [source] Mechanical Surface Properties of CoCr Alloys After Nitrogen PIIIPLASMA PROCESSES AND POLYMERS, Issue S1 2007Inga-Maria Eichentopf Abstract Initial experiments were performed to improve the mechanical surface properties of CoCr alloys by nitrogen plasma immersion ion implantation (PIII). Fast thermally activated diffusion with an activation energy of 0.6 eV was observed. A detailed investigation of the phase formation by X-ray diffraction revealed a variable phase composition with CrN and Cr2N being dominating at higher temperatures and additional fcc phases being present. The hardness of the surface layer increased from about 300 HV by a factor of 3,5 while the wear rate was reduced by a factor of up to ten as a result of the PIII treatment. [source] FK506 and Sildenafil Promote Erectile Function Recovery after Cavernous Nerve Injury Through Antioxidative MechanismsTHE JOURNAL OF SEXUAL MEDICINE, Issue 4i 2007Gwen Lagoda MS ABSTRACT Introduction., Immunophilin ligands and phosphodiesterase type 5 (PDE5) inhibitors are touted to promote erectile function recovery after cavernous nerve (CN) injury. However, the mechanisms for their effects remain unclear. Aim., To compare the erection recovery effects of the immunophilin ligand FK506 and the PDE5 inhibitor sildenafil after CN injury and determine whether they involve antioxidative and/or antiapoptotic mechanisms. Methods., Initial experiments established conditions of our CN injury model in adult male Sprague-Dawley rats. Subsequently, we evaluated treatment effects 14 days after: (i) unilateral CN injury (UNI) + saline (vehicle control); (ii) UNI + FK506 (5 mg/kg once daily, subcutaneous ×5 days); (iii) UNI + sildenafil (20 mg/kg every 8 hours, subcutaneous ×7 days); (iv) UNI + FK506/sildenafil; and (v) sham surgery. Main Outcome Measures., Intracavernous pressure (ICP) measurement after CN electrical stimulation to assess erectile function and Western blot analysis of expressions of glutathione peroxidase (GPX; antioxidant enzyme), nitrotyrosine (NT; oxidative stress marker), and phosphorylated and total Akt (antiapoptotic factor) in penes. Results., In the UNI model, GPX expression was increased at Days 1 and 7, while p-Akt expression decreased at Day 1 and returned to baseline at Day 7. GPX expression was significantly higher in the UNI + FK506 group compared with the saline-treated group (P < 0.05). ICP increased in all treatment groups compared with that of the saline-treated group (P < 0.05). NT levels were increased after saline treatment (P < 0.05) but not after FK506 and sildenafil treatment, alone or in combination. GPX was localized to nerves coursing through the penis and to smooth muscle and endothelium of the dorsal vein and arteries. Conclusions., Both FK506 and sildenafil protect erectile function after CN injury by decreasing oxidative stress-associated tissue damage. FK506 may act through increased GPX activity. Further research is required to elucidate mechanisms associated with the beneficial effect of sildenafil. Lagoda G, Jin L, Lehrfeld TJ, Liu T, and Burnett AL. FK506 and sildenafil promote erectile function recovery after cavernous nerve injury through antioxidative mechanisms. J Sex Med 2007;4:908,916. [source] Assessment of Intraoperative Safety in Transoral Robotic Surgery,THE LARYNGOSCOPE, Issue 2 2006Neil G. Hockstein MD Abstract Introduction: Robotic technology has been safely integrated into thoracic and abdominopelvic surgery, and the early experience has been very promising with very rare complications related to robotic device failure. Recently, several reports have documented the technical feasibility of transoral robotic surgery (TORS) with the daVinci Surgical System. Proposed pharyngeal and laryngeal applications include radical tonsillectomy, base-of-tongue resection, supraglottic laryngectomy, and phonomicrosurgery. The safety of transoral placement of the robotic endoscope and instruments has not been established. Potential risks specific to the transoral use of the surgical robot include facial skin laceration, tooth injury, mucosal laceration, mandible fracture, cervical spine fracture, and ocular injury. We hypothesize that these particular risks of transoral surgery are similar with robotic assistance compared with conventional transoral surgery. Methods: To test this hypothesis, we attempted to intentionally injure a human cadaver with the daVinci Surgical System by impaling the facial skin and pharyngeal and laryngeal mucosa with the robotic instruments and endoscope. We also attempted to extract or fracture teeth and fracture the cadaver's mandible and cervical spine by applying maximal pressure and torque with the robotic arms. Experiments were documented with still and video photography. Results: Impaling the cadaver's skin and mucosa resulted in only superficial lacerations. Tooth, mandible, and cervical spine fracture could not be achieved. Conclusions: Initial experiments performing TORS on a human cadaver with the daVinci Surgical System demonstrate a safety profile similar to conventional transoral surgery. Additionally, we discuss several strategies to increase patient safety in TORS. [source] Functional Impairment of Renal Organic Cation Transport in Experimental DiabetesBASIC AND CLINICAL PHARMACOLOGY & TOXICOLOGY, Issue 4 2002Brett Grover The experiments compared the ability of renal cortex slices from streptozotocin-induced diabetic and non-diabetic rats to accumulate the model cation, 14C-tetraethylammonium under controlled conditions. Initial experiments demonstrated a progressive decline in tetraethylammonium accumulation with increasing duration of diabetes. The maximal decrease was observed at 21 days after streptozotocin injection. Time-dependent incubations revealed that tetraethylammonium uptake from both diabetic and non-diabetic rats followed a curvilinear pattern expected of an active process. However, at steady state the diabetic-derived slices accumulated a significant 38% less tetraethylammonium versus slices from non-diabetics. Concentration-dependent incubations of tetraethylammonium (0.01,10 mM, 60 min.) demonstrated saturable transport in both diabetic and non-diabetic slices with a significantly decreased capacity of diabetic-derived slices to accumulate tetraethylammonium. Cellular respiration rates in the two groups were not different. Insulin treatment of the diabetic rats prevented the transport decline. While the causative factor of the transport impairment in diabetes is unresolved, this study documents an aspect of diabetic nephropathy that has not been previously reported but which may have important implications for renal excretion of cationic drugs and toxicants. The results also provide a mechanism for the well-documented "protection phenomenon" by which the kidneys of diabetic rats are resistant to nephrotoxicity induced by the chemotherapeutic agent cisplatin. [source] Lack of effectiveness of botulinum neurotoxin A on isolated detrusor strips and whole bladders from mice and guinea-pigs in vitroBJU INTERNATIONAL, Issue 10 2009Sarah Howles OBJECTIVE To differentiate between the effects of parasympathetic and sensorimotor stimulation of isolated mouse and guinea-pig bladders in vitro by measuring the pressure increases to electrical field stimulation (EFS) and then comparing the effects of botulinum neurotoxin A (BoNT-A) applied either to the lumen or to the external bathing medium. MATERIALS AND METHODS Isolated mouse and guinea-pig bladders and detrusor strips were exposed to EFS in vitro before and after the addition of BoNT-A. The rationale of this method was that BoNT-A applied to the outside of the bladder would first affect the parasympathetic nerves before diffusing inwards to affect the sensorimotor innervation. BoNT-A applied intravesically would first reach the sensorimotor nerves and only later the parasympathetic nerves. Initial experiments on strips of detrusor were conducted to establish the correct dosage and application time of BoNT-A. RESULTS Contrary to our expectations, BoNT-A application failed to produce any significant effects on either the detrusor strips or whole bladders. CONCLUSIONS Our experimental design failed to show any effect of BoNT-A on the contractility of detrusor muscle strips or whole bladders from mice and guinea-pigs. The reason for this is unclear, but may be related to tissue spending inadequate time incubated with BoNT-A under physiological conditions. [source] Natural-gradient tracer experiments in epikarst: a test study in the Acqua dei Faggi experimental site, southern ItalyGEOFLUIDS (ELECTRONIC), Issue 3 2008E. PETRELLA Abstract Two natural-gradient tracer experiments were carried out using borehole fluorometers in order to characterize the internal structure of epikarstic horizons and analyze subsurface flow within these high-conductivity layers. The experiments were carried out in a test site in southern Italy where the epikarst is made up of an upper part with pervasive karstification and a lower part without pervasive karstification. Injection and observation boreholes were 6.9 m apart. An initial experiment demonstrated that wider (conduits) and narrower (fractures and bedding planes) openings coexist in a well-connected network within the lower epikarst. The adjusted aperture of the opening network (105 ,m) suggests that conduits are subordinately developed. The lower epikarstic horizon is hydraulically similar to granular porous media and Darcy's law can be applied to describe groundwater flow. A small value of longitudinal dispersivity (0.13 m) shows that variations in the velocity field in the direction of flow are less significant than those typical of carbonate systems at the same experiment scale. A second experiment demonstrated that longitudinal dispersivity (2.42 m) in the upper epikarst is in agreement with findings in other carbonates at the same experiment scale. However, despite the higher dispersivity and more pervasive karstification, the mean tracer velocity (3.7 m day,1) in the upper epikarst is slightly lower than the velocity in the lower epikarst (13.6 m day,1). [source] Development of a multifunctional sales response model with the diagnostic aid of artificial neural networksJOURNAL OF FORECASTING, Issue 7 2005Stefania Pantelidaki Abstract This paper proposes an approach that models and forecasts sales through a flexible parametric response function (multifunctional), allowing for differentiated behavioural assumptions of the response determinants to be specified, and uses neural network modelling as a re-specification tool for the response model in order to improve forecasting performance. An initial experiment on a sample of sales data demonstrates feasibility and gives comparative insights via alternative model specifications. Copyright © 2005 John Wiley & Sons, Ltd. [source] Blockade of the 5-HT3 receptor for days causes sustained relief from mechanical allodynia following spinal cord injuryJOURNAL OF NEUROSCIENCE RESEARCH, Issue 2 2009Yuhua Chen Abstract Chronic neuropathic pain is a frequent, serious outcome of spinal cord injury (SCI) that is highly refractory to treatment. Serotonin can contribute to neuropathic pain after SCI, as suggested by our previous observation that transient blockade of the 5-HT3 receptor by intrathecal injections of the antagonist ondansetron reduces mechanical allodynia after SCI in rats. The current study determined whether intrathecal or intravenous infusion of ondansetron for 3 or 7 days, respectively, could cause sustained blockade of mechanical allodynia at and below the level of a twelfth thoracic clip compression injury in rats. Intrathecal 3-day infusion of ondansetron (2.0 ,g/hr), targeted to the cord rostral to the SCI and commencing at 28 days after SCI, decreased at-level mechanical allodynia by 40% and below-level allodynia by 60% compared with saline-treated rats (controls). This reduction was sustained throughout drug delivery and for 1 day afterward. During the next 3 days, allodynia gradually returned toward the values of saline-treated rats. An initial experiment showed that bolus intravenous injections of ondansetron (20,100 ,g) at 28 days after SCI decreased both at- and below-level allodynia for 90,120 min. Intravenous 7-day infusions (20 ,g/hr), commencing at 28 days after SCI, significantly decreased at-level allodynia by 48% and below-level allodynia by 51% compared with controls. This reduction of allodynia lasted throughout the infusion and for 1,3 days afterward while pain responses gradually approached those of controls. These findings suggest a potential role of 5-HT3 receptor antagonism in the relief of neuropathic pain after SCI in humans. © 2008 Wiley-Liss, Inc. [source] ,-Dystroglycan is essential for the induction of Egr3, a transcription factor important in muscle spindle formationDEVELOPMENTAL NEUROBIOLOGY, Issue 7 2010Stacey Williams Abstract Muscle spindle fibers are specialized stretch receptors that allow the perception and coordination of limb movement. The differentiation of these specialized structures is initiated by signals derived from the in growing Ia sensory neurons during development. While the direct molecular signaling mechanisms between sensory neurons and developing muscle at nascent spindle fibers have been well documented in past studies the roles of muscle basal lamina components on this process have not previously been described. As such, our initial experiments addressed potential roles for agrin (AGRN) and laminin (LN) in the expression of the transcription factor Egr3. Levels of Egr3 were monitored using immunoblot analysis and both basal lamina molecules proved effective in inducing Erg3 expression. Previous work had established neuregulin (NRG) as a critical signaling component in spindle fiber development so blocking experiments with NRG and ErbB inhibitors were then used to determine if LN-induced Egr3 expression was occurring as a result of NRG-ErbB signaling and not via other, novel pathway. Inhibiting signaling through this pathway did indeed reduce the expression of Egr3. Finally, we looked at ,-dystrogylcan, a shared receptor for AGRN and LN at neuromuscular junctions. Using a ,-dystroglycan (,-DG) silenced muscle cell line and an anti-,-DG antibody we attempted to block basal lamina/,-DG interactions. Again, and in both instances, Egr3 expression was significantly decreased. Taken together, analysis of the results from these experiments revealed that indeed AGRN, LN, and ,-DG influence Egr3 levels and therefore may play an important role in spindle fiber differentiation. © 2010 Wiley Periodicals, Inc. Develop Neurobiol 70:498,507, 2010 [source] The Langerhans' cell-like cell lines XS52 and XS106 express mRNA for ciliary neurotrophic factor and neurotrophic factor 4/5EXPERIMENTAL DERMATOLOGY, Issue 9 2004K. Seiffert Neurotrophins are responsible for the survival and outgrowth of nerves within the peripheral and central nervous systems. These factors include brain-derived neurotrophic factor (BDNF), CNTF, NT 3, and NT4/5. We have previously shown that LCs lie in close proximity to nerves and that several neuropeptides regulate LC function, implying that nerves send regulatory signals to LCs. To evaluate the possibility that LC signal nerves by release of neurotrophins, we examined LC expression of neurotrophins by RT-PCR. To eliminate the possibility of contaminating keratinocytes in highly enriched LC preparations, we utilized the LC-like cell lines XS52 (BALB/c derived) and XS106 (A/J derived) for initial experiments. The RNA obtained was digested with DNase to ensure complete absence of genomic DNA. Several independent RT-PCRs revealed expression of bands of the expected size for CTNF and NT4/5, but not for BDNF and NT3 in XS106 and XS52 cells. In contrast, the transformed keratinocyte cell line PAM212 expressed BDNF, as well as CTNF and NT4/5. Preliminary experiments with purified LC confirm the expression of CTNF and NT4/5 and also show the expression of BDNF. However, we cannot be sure that BDNF expression is not due to keratinocyte contamination. We conclude that LCs may regulate nerve cells by the release of neurotrophic factors. [source] The Role of Verbal Repetition in the Development of Infant Speech Preferences From 4 to 14 Months of AgeINFANCY, Issue 2 2009Gerald W. McRoberts Four experiments investigated infants' preferences for age-appropriate and age-inappropriate infant-directed speech (IDS) over adult-directed speech (ADS). Two initial experiments showed that 6-, 10-, and 14-month-olds preferred IDS directed toward younger infants, and 4-, 8-, 10-, and 14-month-olds, but not 6-month-olds, preferred IDS directed toward older infants. In Experiment 3. 6-month-olds preferred IDS directed toward older infants when the frequency of repeated utterances matched IDS to younger infants. In Experiment 4, 6-month-olds preferred repeated IDS utterances over the same IDS utterances organized without repetition. Attention to repeated utterances precedes word segmentation and sensitivity to statistical cues in continuous speech, and might play a role in the discovery of these and other aspects of linguistic structure. [source] Efficient synthesis of ethanol and acetic acid from methane and carbon dioxide with a continuous, stepwise reactorAICHE JOURNAL, Issue 5 2010W. Huang Abstract The synthesis of C2 -oxygenates such as ethanol and acetic acid accomplished by CH4 dissociation and subsequent CO2 insertion onto methyl radicals, named the stepwise reaction technology, has been demonstrated to be both feasible and efficient through initial experiments conducted in microreactor units. This article describes the development of this technology, highlighting the aforementioned stepwise technology using a dual-reactor system, which can ensure that two raw gases enter the reactor uninterruptedly and are not mixed after reaction. The system productivity for acetic acid and ethanol displayed efficiencies greater than 5,10 times that of previous microreactor units. The investigation of mechanism indicates that acetic acid arises from insertion of CO2 into MCHx, while ethanol is formed either by hydrogenation of acetic acid or by hydration of C2H4, which results from homo-coupling of CH4. The latter route is the preferred of the two. © 2009 American Institute of Chemical Engineers AIChE J, 2010 [source] Design and use of multi-affinity surfaces in biomolecular interaction analysis,mass spectrometry (BIA/MS): a step toward the design of SPR/MS arraysJOURNAL OF MOLECULAR RECOGNITION, Issue 1 2003Dobrin Nedelkov Abstract The feasibility of multi-affinity ligand surfaces in biomolecular interaction analysis,mass spectrometry (BIA/MS) was explored in this work. Multi-protein affinity surfaces were constructed by utilizing antibodies to beta-2-microglobulin, cystatin C, retinol binding protein, transthyretin, serum amyloid P and C-reactive protein. In the initial experiments, all six antibodies were immobilized on a single site (flow cell) on the sensor chip surface, followed by verification of the surface activity via separate injections of purified proteins. After an injection of diluted human plasma aliquot over the antibodies-derivatized surfaces, and subsequent MALDI-TOF MS analysis, signals representing five out of the six targeted proteins were observed in the mass spectra. Further, to avoid the complexity of the spectra, the six proteins were divided into two groups (according to their molecular weight) and immobilized on two separate surfaces on a single sensor chip, followed by an injection of human plasma aliquot. The resulting mass spectra showed signals from all proteins. Also, the convolution resulting from the multiply charged ion species was eliminated. The ability to create such multi-affinity surfaces indicates that smaller-size ligand areas/spots can be employed in the BIA/MS protein interaction screening experiments, and opens up the possibilities for construction of novel multi-arrayed SPR-MS platforms and methods for high-throughput parallel protein interaction investigations. Copyright © 2003 John Wiley & Sons, Ltd. [source] Optimizing DNA yield from buccal swabs in the elderly: Attempts to promote buccal cell growth in cultureAMERICAN JOURNAL OF HUMAN BIOLOGY, Issue 5 2003David J. Vandenbergh Participation in genetic studies is often limited by a volunteer's reluctance to donate blood samples. We wished to determine if alternate, less painful, methods to venipuncture could be used to collect cells to provide DNA for genotyping, and whether the cells could be grown in culture for extraction of DNA. Volunteers in the study were comprised of two groups. Nine individuals from a university campus were recruited to provide samples for initial experiments. A second group of 710 twins and singletons from North Carolina and of African-American descent were a part of an ongoing study of age-related traits and participated in collection of buccal swabs via the mail. A protocol was generated that maximizes the recovery of DNA from buccal swabs, which are easier to handle than saline rinses. The DNA recovered is stable over several years, allowing genotype tests at a future date. Attempts to encourage growth of buccal epithelial cells recovered from swabs in tissue culture proved unsuccessful. Buccal swabs work well for the collection of DNA, especially from nonclinic-based volunteers, and can be sent via the mail to the laboratory for DNA extraction. Thus, an inexpensive and efficient method exists for genetic studies of population-based samples. Am. J. Hum. Biol. 15:637,642, 2003. © 2003 Wiley-Liss, Inc. [source] Using gene chips to identify organ-specific, smooth muscle responses to experimental diabetes: potential applications to urological diseasesBJU INTERNATIONAL, Issue 2 2007Jason D. Hipp OBJECTIVE To identify early diabetes-related alterations in gene expression in bladder and erectile tissue that would provide novel diagnostic and therapeutic treatment targets to prevent, delay or ameliorate the ensuing bladder and erectile dysfunction. MATERIALS AND METHODS The RG-U34A rat GeneChip® (Affymetrix Inc., Sunnyvale, CA, USA) oligonucleotide microarray (containing ,8799 genes) was used to evaluate gene expression in corporal and male bladder tissue excised from rats 1 week after confirmation of a diabetic state, but before demonstrable changes in organ function in vivo. A conservative analytical approach was used to detect alterations in gene expression, and gene ontology (GO) classifications were used to identify biological themes/pathways involved in the aetiology of the organ dysfunction. RESULTS In all, 320 and 313 genes were differentially expressed in bladder and corporal tissue, respectively. GO analysis in bladder tissue showed prominent increases in biological pathways involved in cell proliferation, metabolism, actin cytoskeleton and myosin, as well as decreases in cell motility, and regulation of muscle contraction. GO analysis in corpora showed increases in pathways related to ion channel transport and ion channel activity, while there were decreases in collagen I and actin genes. CONCLUSIONS The changes in gene expression in these initial experiments are consistent with the pathophysiological characteristics of the bladder and erectile dysfunction seen later in the diabetic disease process. Thus, the observed changes in gene expression might be harbingers or biomarkers of impending organ dysfunction, and could provide useful diagnostic and therapeutic targets for a variety of progressive urological diseases/conditions (i.e. lower urinary tract symptoms related to benign prostatic hyperplasia, erectile dysfunction, etc.). [source] | |||||||||||||||||||