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D L Harris

Publications and source records attributed to D L Harris.

At least 37 records · Page 2Linked to original sources
PubMed2000

Determinants of recognition of ligands binding to benzodiazepine receptor/GABA(A) receptors initiating sedation.

Complementary behavioral and computational studies of 21 structurally diverse, gamma-amino butyric acid (GABA)(A) benzodiazepine receptor ligands that influence spontaneous locomotor activity have been performed in this work. This behavioral endpoint is a well-accepted indicator of sedation particularly for GABA(A)/benzodiazepine receptor ligands. The goal of the work presented here is the identification and assessment of the minimum requirements for ligand recognition of GABA(A)/benzodiazepine receptors leading to activity at the sedation endpoint embedded in a common 3D pharmacophore for recognition. Using the experimental results, together with a systematic computational procedure developed in our laboratory, a five-component 3D pharmacophore for recognition of the GABA(A) receptor subtypes associated with the sedative behavioral response has been developed consisting of: two proton-accepting moieties, a hydrophobic region, a ring with polar moieties and an aromatic ring in a common geometric arrangement in all ligands having an effect at the sedation endpoint. To provide further evidence that the 3D pharmacophore developed embodied common requirements for receptor recognition, a pharmacophore analysis was performed for agonists, inverse agonists and antagonists separately. Each of the resulting pharmacophores contained the same five moieties at comparable distances to those found for the pharmacophore generated using all of them together. This result confirms that this pharmacophore constitutes a recognition pharmacophore representing required features in the overlapping portion of their binding sites. The reliability of this 3D pharmacophore was then assessed in several ways. First, it was determined that ligands that had no effect at the sedation endpoint did not comply with the pharmacophore requirements. Second, four benzodiazepine receptor ligands known to have an effect at the sedation endpoint, but not used in the pharmacophore development were found to satisfy the requirements of this pharmacophore. Third, the geometric and chemical requirements embedded in this pharmacophore were used to search 3D databases resulting in the identification of benzodiazepine receptor ligands known to affect sedation, but not included in the pharmacophore development. Finally, a 3D-quantitative structure analysis procedure (QSAR) model was developed based upon the ligands in the training set superimposed at their sedation pharmacophore points. The 3D-QSAR model shows good predictivity for binding of these ligands to receptor subtypes containing alpha1 but not alpha5 subunits. The pharmacophore developed for the sedation endpoint thus provides a predictive binding model for diverse ligand binding to alpha1 containing receptor subtypes.

Animals↗
PubMed2000

Development of a 3D pharmacophore for nonspecific ligand recognition of alpha1, alpha2, alpha3, alpha5, and alpha6 containing GABA(A)/benzodiazepine receptors.

Transfected cells containing GABA(A) benzodiazepine receptors (BDZRs) have been utilized to systematically determine the affinity of ligands at alpha1, alpha2, alpha3, alpha5 and alpha6 subtypes in combination with beta2 and gamma2. All but a few of the ligands thus far studied have relatively high affinities for each of these alpha subtype receptors. Thus, these ligands must contain common stereochemical properties favorable for recognition by each of the subtype combinations. In the present work, such a common three-dimensional (3D) pharmacophore for recognition of alpha1, alpha2, alpha3, alpha5 and alpha6 containing GABA(A)/BDZRs types of receptors has been developed and assessed, using as a database receptor affinities measured in transfected cells for 27 diverse compounds. The 3D-recognition pharmacophore developed consists of three proton accepting groups, a hydrophobic group, and the centroid of an aromatic ring found in a common geometric arrangement in the 19 nonselective ligands used. Three tests were made to assess this pharmacophore: (i) Four low affinity compounds were used as negative controls, (ii) Four high affinity compounds, excluded from the pharmacophore development, were used as compounds for pharmacophore validation, (iii) The 3D pharmacophore was used to search 3D databases. The results of each of these types of assessments provided robust validation of the 3D pharmacophore. This 3D pharmacophore can now be used to discover novel nonselective ligands that could be activation selective at different behavioral end points. Additionally, it may serve as a guide in the design of more selective ligands, by determining if candidate ligands proposed for synthesis conform to this pharmacophore and selecting those that do not for further experimental assessment.

Benzodiazepines↗
PubMed2000

Development and assessment of a 3D pharmacophore for ligand recognition of BDZR/GABAA receptors initiating the anxiolytic response.

Benzodiazepine receptor (BDZR) ligands are structurally diverse compounds that bind to specific binding sites on GABAA receptors and allosterically modulate the effect of GABA on chloride flux. The binding of BDZR ligands to this receptor system results in activity at multiple behavioral end points including anxiolytic, sedative, hyperphagic, anticonvulsant and hyperthermic effects. In the work presented here, 17 structurally diverse BDZR ligands of the receptors initiating the anxiolytic response have been studied using a systematic computational procedure developed in our laboratory. Using this procedure, a five component 3D recognition pharmacophore was obtained consisting of two proton acceptors, a hydrophobic group, an aromatic electron accepting ring and a ring containing polar moieties, all found in a common geometric arrangement in the 15 compounds with an effect at the anxiolytic end point and absent in two control compounds. The 3D pharmacophore developed was validated by searching 3D databases and finding known BDZR ligands active at the anxiolytic end point, including 1,4-BDZ derivatives, imidazo BDZ and beta-carboline ligands.

Anti-Anxiety Agents↗
PubMed2000

An approach to the control of disease transmission in pig-to-human xenotransplantation.

Although several major immunologic hurdles need to be overcome, the pig is currently considered the most likely source animal of cells, tissues and organs for transplantation into humans. Concerns have been raised with regard to the potential for the transfer of infectious agents with the transplanted organ to the human recipient. This risk is perceived to be increased as it is likely that the patient will be iatrogenically immunocompromised and the organ-source pig may be genetically engineered in such a way to render its organs particularly susceptible to infection with human viruses. Furthermore, the risk may not be restricted to the recipient, but may have consequences for the health of others in the community. The identification of porcine endogenous retroviruses and of hitherto unknown viruses have given rise to the most concern. We document here the agents we believe should be excluded from the organ-source pigs. We discuss the likelihood of achieving this aim and outline the potential means by which it may best be achieved.

Animals↗
PubMed2000

Benzodiazepine-induced hyperphagia: development and assessment of a 3D pharmacophore by computational methods.

Benzodiazepine receptor (BDZR) ligands are structurally diverse compounds that bind to specific binding sites on GABA(A) receptors and allosterically modulate the effect of GABA on chloride ion flux. The binding of BDZR ligands to this receptor system results in activity at multiple behavioral endpoints, including anxiolytic, sedative, anticonvulsant, and hyperphagic effects. In the work presented here, a computational procedure developed in our laboratory has been used to obtain a 3D pharmacophore for ligand recognition of the GABA(A)/BDZRs initiating the hyperphagic response. To accomplish this goal, 17 structurally diverse compounds, previously assessed in our laboratory for activity at the hyperphagic endpoint, were used. The result is a four-component 3D pharmacophore. It consists of two proton acceptor atoms, the centroid of an aromatic ring and the centroid of a hydrophobic moiety in a common geometric arrangement in all compounds with activity at this endpoint. This 3D pharmacophore was then assessed and successfully validated using three different tests. First, two BDZR ligands, which were included as negative controls in the set of seventeen compounds used for the pharmacophore development, did not fit the pharmacophore. Second, some benzodiazepine ligands known to have activity at the hyperphagia endpoint, but not included in the pharmacophore development, were used as positive controls and were found to fit the pharmacophore. Finally, using the 3D pharmacophore developed in the present work to search 3D databases, over 50 classical benzodiazepines were found. Among them, were benzodiazepine ligands known to have an effect at the hyperphagic endpoint. In addition, the novel compounds also found in this search are promising therapeutic agents that could beneficially affect feeding behavior.

Benzodiazepines↗
PubMed2000

Weight loss, not aerobic exercise, improves pulmonary function in older obese men.

BACKGROUND: We evaluated the effect of weight loss (WL) or aerobic exercise (AEX) on pulmonary function in middle-aged and older (46-80 years) obese, sedentary men to determine the effect of reductions in body weight and increases in cardiorespiratory fitness on pulmonary function. METHODS: Subjects were randomly assigned to WL (n = 73), AEX (n = 71), or control (n = 26) groups. Maximal oxygen uptake (VO2max), body composition and anthropometrics, pulmonary function, and arterial blood gases were measured at baseline and after interventions. RESULTS: The 35 subjects who completed WL decreased weight by 11%, body fat percentage by 21% (p < .001), waist circumference by 8%, waist-hip ratio by 2%, and fat-free mass by 3% (p < .05). This resulted in a 3% increase in forced vital capacity (FVC) (4.08 +/- 0.71 L vs 4.21 +/- 0.76 L), a 5% increase in total lung capacity (6.62 +/- 0.99 L vs 6.94 +/- 0.99 L), an 18% increase in functional residual capacity (3.09 +/- 0.58 L vs 3.66 +/- 0.79 L), and an 8% increase in residual volume (2.20 +/- 0.44 L vs 2.37 +/- 0.52 L), with no change in forced expiratory volume in one second (FEV1), FEV1/FVC ratio, or carbon monoxide diffusing capacity. The change in FVC correlated with change in body weight (r = -.34, p < .05). The 38 subjects who completed AEX increased VO2max by 14%, with no change in pulmonary function. There were no changes in 8 control subjects. CONCLUSIONS: WL changes static lung volumes, not dynamic pulmonary function, in middle-aged and older, moderately obese, sedentary men. Some of the alterations in static lung function associated with aging may be due to the development of obesity and are modifiable by WL.

Age Factors↗
PubMed1999

Balanitis xerotica obliterans complicating hypospadias repair.

We review the literature and report a series of eight cases of balanitis xerotica obliterans (BXO) developing in patients following primary hypospadias repair. The ages of these patients ranged from 8 to 25 years with BXO developing from 1 to 16 years postoperatively. Six patients were treated by excision of the BXO tissue and two-stage urethroplasties with full-thickness grafts. Three of these patients had further recurrence of BXO and had re-do urethroplasty using a combination of bladder and buccal mucosa. The last two patients in this series had re-do urethroplasty using bladder mucosa only and bladder-buccal mucosa technique, respectively, as first choice for BXO complicating their hypospadias repair.

Adolescent↗
PubMed1999

Transforming growth factor-beta suppresses proliferation of rabbit corneal endothelial cells in vitro.

Corneal endothelial cells in vivo appear to be inhibited in G1 phase of the cell cycle. Studies were carried out to determine whether cultured rabbit corneal endothelium expresses transforming growth factor-beta (TGF-beta) receptor types I, II, and III, suggesting they would be sensitive to a TGF-beta-induced signal. In addition, we explored if TGF-beta might mediate this G1 phase inhibition by implementing flow cytometry and 5-bromo-2'-deoxyuridine (BrdU) immunofluorescence. Reverse transcription-polymerase chain reaction (RT-PCR) products of the expected size were obtained for all three TGF-beta receptor types. Flow cytometry revealed a dose-dependent suppression in the percentage of S phase cells in cultures treated with TGF-beta1 or TGF-beta2. The lowest percentage of S phase cells was found for 10 ng/ml TGF-beta1 and 0.1 ng/ml TGF-beta2. BrdU, an S phase marker, was immunolocalized, and semiquantitative analysis of stained cells showed a maximum suppression of S phase entry at 18 h for 10 ng/ml of TGF-beta11 and 24 h for 10 ng/ml of TGF-beta2. In rabbit, the corneal endothelium expresses TGF-beta receptor types I, II, and III, permitting a TGF-beta signal to be transduced. Flow cytometry reveals a dose-dependent response to both TGF-beta1 and TGF-beta2, and the cells are more sensitive to TGF-beta2. At optimal TGF-beta concentrations, the percentage of S phase cells is comparable to that of a non-proliferating culture, suggesting TGF-beta prevents the cells from proceeding through the G1/S phase transition. This suppression was also seen with BrdU labeling. Together, these results indicate that TGF-beta could be one of the pathways that leads to G1 phase arrest in corneal endothelial cells.

Animals↗
PubMed1999

When (not if) evaluation flexibility is desirable. Examples from the CPHPE initiative.

The evaluation literature often debates whether evaluators should be flexible in evaluation design and activities in order to collaborate with program directors and be responsive to programming needs. Two conditions are specified under which evaluation flexibility is not only desirable but essential. Two examples from the cluster evaluation of the W. K. Kellogg Foundation's Community Partnerships for Health Professions Education initiative are provided to illustrate why flexibility under these conditions proved to be essential. One of the examples, related to the "community" involvement in the initiative, illustrates the need for flexibility as programs experience goals clarification. The other example, related to the coincidental national health care reform efforts, illustrates the need for flexibility both to capture programs' efforts to protect their integrity and to ensure against spurious conclusions as a result of external turbulence in policy environments. How the cluster evaluation team addressed these issues is also described.

Allied Health Personnel↗
PubMed1999

TGF-beta2 in aqueous humor suppresses S-phase entry in cultured corneal endothelial cells.

PURPOSE: Corneal endothelium in vivo is arrested in G1, the phase of the cell cycle that prepares cells for DNA synthesis. In many cell types, transforming factor (TGF)-beta inhibits proliferation by inducing G1-phase arrest. Evidence indicates that corneal endothelial cells synthesize mRNA for TGF-beta1 and are also bathed in aqueous humor that contains TGF-beta2 (mainly in a latent form). As such, this cytokine may maintain the corneal endothelium in a G1-phase-arrested state in vivo. The purpose of these studies was to determine the effect of exogenous TGF-beta2 and TGF-beta2 in aqueous humor on DNA synthesis in cultured corneal endothelial cells. METHODS: Rat corneal endothelial cells were grown in explant culture and identified by morphology and reverse transcription-polymerase chain reaction using primers for specific corneal cell markers. Subconfluent cells were synchronized in the G0 phase (quiescence) by serum starvation for 24 hours. Serum was then added to the cells in the presence or absence of exogenous TGF-beta2 or activated rat aqueous humor. [3H]Thymidine was added, and radioactivity was measured at various time points to detect DNA synthesis. Preincubation of exogenous TGF-beta2 or activated rat aqueous humor with neutralizing antibody was used to test for cytokine specificity. RESULTS: A linear increase in [3H]thymidine incorporation began approximately 16 hours after serum addition, and peak incorporation occurred at approximately 24 hours. Exposure of cells to serum plus TGF-beta2 suppressed [3H]thymidine incorporation in a dose-dependent manner at concentrations ranging from 5 pg/ml to 5 ng/ml. [3H]Thymidine incorporation was also suppressed in cells exposed to serum plus rat aqueous humor diluted 1:10. Neutralizing antibody reversed the effects of both exogenous TGF-beta2 and aqueous humor. CONCLUSIONS: Exogenous TGF-beta2 and TGF-beta2 in aqueous humor suppress S-phase entry of rat corneal endothelial cells. These results suggest that this cytokine in aqueous humor could help maintain the corneal endothelium in a G1-phase-arrested state in vivo.

Animals↗
PubMed1998

Multiplex PCR assay for detection of Clostridium perfringens in feces and intestinal contents of pigs and in swine feed.

A multiplex polymerase chain reaction (PCR) assay, developed to detect the alpha-toxin and enterotoxin genes (cpa and cpe, respectively) of Clostridium perfringens, was used to identify enterotoxigenic isolates of this organism from feces and intestinal contents of pigs and from feed samples from pig farms in Iowa. The organism was grown on tryptose-sulfite-cycloserine (TSC) agar, TSC agar without egg-yolk, sheep blood agar, or in brain heart infusion broth or cooked meat medium. DNA was extracted by boiling and the PCR assay was carried out using reagents from a commercial kit. The 319 bp amplification product of cpa and the 364 bp product of cpe were visualized under UV light after electrophoresis in a 2% agarose gel containing ethidium bromide. The average sensitivity of the assay, determined on artificially contaminated feces, was 9.2 x 10(4) colony forming units per gram. Assay of 97 isolates from feces and intestinal contents revealed cpa in 89, but all were negative for cpe. While 28% of the 442 total samples cultured yielded C. perfringens, only 5% of 298 fecal or intestinal contents samples were positive upon direct examination by the PCR assay. Ninety-one and eight-tenths % of isolates with the phenotype of C. perfringens were cpa positive by PCR. Forty-three percent of feed samples were culture positive, while 48.3% were PCR positive for cpa. None of these were cpe positive. We conclude that PCR is a useful assay for rapid detection of C. perfringens in feed, and for confirmation of the identity of isolates presumed to be C. perfringens.

Animal Feed↗
PubMed1998

Multidisciplinary education outcomes of the W.K. Kellogg Community Partnerships and Health Professions Education initiative.

This evaluation highlights several points to be considered by others instituting multidisciplinary approaches to health professions education. Community-based, multidisciplinary experiences appear to reinforce support students' interests in pursuing primary care careers. The multidisciplinary, community-based approach to health professions education did not affect academic learning. Project leaders and students reported no risks in terms of board scores between CPHPE students and others in traditional programs. The multidisciplinary, community-based approach to health professions education created opportunities at some sites for students to see "team medicine" in action. It was practical and helped students to understand how they could be more effective in their roles as opposed to competitive. Students require socialization within their own individual disciplines as well as socialization across disciplines. The differences in the structures of traditional health professions education schools interfered with the development of multidisciplinary contexts for learning at some sites. Campus faculty were not necessarily socialized to engage in multidisciplinary efforts. Their disciplines generally do not recognize and reward this behavior. Early and continuous faculty development may significantly help projects to improve communication and develop a better understanding of the contexts of curricular changes across disciplines. This evaluation was exploratory. Further research is needed to better understand what forms of multidisciplinary curriculum are most effective and economically feasible, what forms survive over time, and whether the intended final outcomes of the CPHPE initiative are achieved, not only with medical students but also with the other health professions students.

Curriculum↗
PubMed1998

Livestock improvement: art, science, or industry?

The shift in perspectives about livestock improvement over a 70-yr period is reviewed. The view changes from livestock improvement as an art, through its becoming a science, to its application as a technology in industry. The use of scientific tools in corporate settings for poultry, dairy, and swine breeding usually involves a strong focus on the economics of production, and this focus determines that selection indexes should be used. Such focus has seemed to not have been present in breeding for the extensively managed species and livestock classes but seems needed in order to provide a basis for organization and direction of positive contributions of the breeding segment to the production segment.

Animal Husbandry↗
PubMed1998

Evaluation of antemortem polymerase chain reaction and serologic methods for detection of Lawsonia intracellularis-exposed pigs.

OBJECTIVE: To evaluate polymerase chain reaction (PCR) for detection of Lawsonia intracellularis DNA in feces and an indirect fluorescent antibody test (IFAT) for detecting serum IgG antibodies in pigs exposed to L intracellularis. ANIMALS: 15 seven-week-old pigs and 42 three-week-old pigs. PROCEDURE: During 3 experiments, 23 pigs were inoculated with a pure culture of L intracellularis, 31 pigs served as noninoculated controls, and 3 pigs were used as sentinels. Fecal shedding of L intracellularis was monitored by use of PCR analysis at 7-day intervals. At euthanasia, the ileum was obtained for PCR and histologic analyses. Serum was obtained at 7-day intervals for use in the IFAT. RESULTS: Polymerase chain reaction analysis detected L intracellularis DNA in the feces of 39% of the inoculated pigs; by postinoculation days 21 to 28, 90% of inoculated pigs developed IgG antibodies detected by IFAT. Neither L intracellularis DNA nor IgG antibodies were detected in any of the noninoculated control pigs at euthanasia. Sera from pigs inoculated with enteric pathogens other than L intracellularis did not contain detectable antibodies that reacted with L intracellularis by use of the IFAT. CONCLUSION: The IFAT for L intracellularis IgG antibody detection appeared to be a more sensitive antemortem test for detecting pigs experimentally infected with L intracellularis than was a PCR method for direct detection of the organism in the feces. CLINICAL RELEVANCE: Not all animals that are infected with L intracellularis shed the organism in feces at detectable amounts.

Animals↗
PubMed1998

Mitotic inhibition of corneal endothelium in neonatal rats.

PURPOSE: Corneal endothelium in humans does not divide to any significant extent after birth; therefore, with age there is a gradual loss of cells. When cell density is reduced to a critical level, the endothelium cannot function to maintain corneal clarity, and the cornea becomes permanently cloudy. Currently, the blindness that results can be treated only by corneal transplantation. The long-term goal is to find methods to stimulate corneal endothelial proliferation in a clinically relevant manner. The first step toward achieving this goal is to identify mechanisms responsible for the induction and maintenance of mitotic inhibition of the corneal endothelium in vivo. During corneal development, the endothelium is formed by migration and proliferation of mesenchymal cells from the ocular periphery. Soon after the monolayer is formed, proliferation ceases. In tissue culture, many cell types cease proliferating upon formation of stable cell-cell and cell-substrate attachments. The goal of the present studies was to determine whether establishment of stable contacts correlates with cessation of endothelial proliferation during corneal development in vivo. METHODS: Corneas from neonatal (days 1, 3, 7, 10, 13, 14, 17, 21, 28, and 42) and adult rats were used for immunolocalization of the following: bromodeoxyuridine (BrdU), an S-phase marker; p27kip1 and p21cip1, G1-phase inhibitors; connexin-43 and ZO-1, proteins associated with gap and tight junctions, respectively; Na+/K+-ATPase and beta3-integrin, markers of plasma membrane polarity; and fibronectin and collagen type IV, constituents of Descemet's membrane. Nuclei staining positively for BrdU were counted to determine the relative number of S-phase cells at various times after birth. Marker protein expression and localization were determined by conventional fluorescence microscopy and by confocal microscopy. RESULTS: The number of endothelial cells staining positively for BrdU gradually decreased between postnatal days 1 and 13. After postnatal day 13, positive BrdU staining was no longer detectable. During the first postnatal week, cells stained positively for the G1-phase inhibitor p27kip1 but not for p21cip1. Connexin-43 achieved its mature location by postnatal day 1. ZO-1, Na+/K+-ATPase, beta3-integrin, fibronectin, and collagen type IV achieved their mature localization patterns between postnatal days 14 and 21. CONCLUSIONS: In neonatal rat, corneal endothelial cells are still entering the cell cycle at birth, but cell cycle entry gradually decreases, so that by postnatal day 13 cells are no longer entering the S-phase. The G1-phase inhibitor p27kip1, but not p21cip1, may help mediate this inhibition. Stable cell-cell and cell-substrate contacts gradually form, and monolayer maturation is complete between postnatal days 14 and 21. The results lead to the hypothesis that, in developing rat cornea in vivo, the establishment of stable cell-cell and cell-substrate contacts initiates a cascade of events, mediated by p27kip1, which induces mitotic inhibition in the endothelial monolayer.

Animals↗