PubMed Health⌕ Search

Biomedical subjects

David L Smith

Publications and source records attributed to David L Smith.

At least 37 records · Page 2Linked to original sources
PubMed2005

An obligatory intermediate controls the folding of the alpha-subunit of tryptophan synthase, a TIM barrel protein.

The proposed kinetic folding mechanism of the alpha-subunit of tryptophan synthase (alphaTS), a TIM barrel protein, displays multiple unfolded and intermediate forms which fold through four parallel pathways to reach the native state. To obtain insight into the secondary structure that stabilizes a set of late, highly populated kinetic intermediates, the refolding of urea-denatured alphaTS from Escherichia coli was monitored by pulse-quench hydrogen exchange mass spectrometry. Following dilution from 8 M urea, the protein was pulse-labeled with deuterium, quenched with acid and mass analyzed by electrospray ionization mass spectrometry (ESI-MS). Hydrogen bonds that form prior to the pulse of deuterium offer protection against exchange and, therefore, retain protons at the relevant amide bonds. Consistent with the proposed refolding model, an intermediate builds up rapidly and decays slowly over the first 100 seconds of folding. ESI-MS analysis of the peptic fragments derived from alphaTS mass-labeled and quenched after two seconds of refolding indicates that the pattern of protection of the backbone amide hydrogens in this transient intermediate is very similar to that observed previously for the equilibrium intermediate of alphaTS highly populated at 3 M urea. The protection observed in a contiguous set of beta-strands and alpha-helices in the N terminus implies a significant role for this sub-domain in directing the folding of this TIM barrel protein.

Escherichia coli↗
PubMed2005

Predictive spatial dynamics and strategic planning for raccoon rabies emergence in Ohio.

Rabies is an important public health concern in North America because of recent epidemics of a rabies virus variant associated with raccoons. The costs associated with surveillance, diagnostic testing, and post-exposure treatment of humans exposed to rabies have fostered coordinated efforts to control rabies spread by distributing an oral rabies vaccine to wild raccoons. Authorities have tried to contain westward expansion of the epidemic front of raccoon-associated rabies via a vaccine corridor established in counties of eastern Ohio, western Pennsylvania, and West Virginia. Although sporadic cases of rabies have been identified in Ohio since oral rabies vaccine distribution in 1998, the first evidence of a significant breach in this vaccine corridor was not detected until 2004 in Lake County, Ohio. Herein, we forecast the spatial spread of rabies in Ohio from this breach using a stochastic spatial model that was first developed for exploratory data analysis in Connecticut and next used to successfully hind-cast wave-front dynamics of rabies spread across New York. The projections, based on expansion from the Lake County breach, are strongly affected by the spread of rabies by rare, but unpredictable long-distance translocation of rabid raccoons; rabies may traverse central Ohio at a rate 2.5-fold greater than previously analyzed wildlife epidemics. Using prior estimates of the impact of local heterogeneities on wave-front propagation and of the time lag between surveillance-based detection of an initial rabies case to full-blown epidemic, specific regions within the state are identified for vaccine delivery and expanded surveillance effort.

Animals↗
PubMed2005

Strategic interactions in multi-institutional epidemics of antibiotic resistance.

The increasing frequency of antibiotic resistance in hospital-acquired infections is a major public health concern that has both biological and economic causes. Here we develop conceptual mathematical models that couple the economic incentives and population biology of hospital infection control (HIC). We show that the optimal investment by a hospital for HIC changes with the proportion of patients already colonized with antibiotic-resistant bacteria (ARB) at the time of admission. As that proportion increases, the optimal behavior of a hospital is to increase spending to control ARB with low transmissibility and decrease spending on those with high transmissibility. In some cases, the global optimum investment in HIC can shift discontinuously from one that contains transmission to a do-nothing policy once the proportion already colonized at the time of admission becomes too great. We also show that investments in HIC are determined by a strategic game when several hospitals share patients. Hospitals acting selfishly and rationally will free-ride on the investments of other hospitals, and the level of free-riding should increase with the number of other hospitals in the area. Thus, in areas with many hospitals, the rational strategy for each hospital is to spend less than in areas with few hospitals. Thus, we predict that transmission rates and the prevalence of ARB should be higher in urban hospitals, for instance, compared with rural hospitals. We conclude that regional coordination and planning for HIC is an essential element of public health planning for hospital-acquired infections.

Cross Infection↗
PubMed2005

A prospective evaluation of ultrasound-directed transparenchymal vascular control with linear cutting staplers in major hepatic resections.

BACKGROUND: We prospectively evaluated a novel ultrasound-directed technique of major hepatic resection using transparenchymal application of vascular staplers intending to minimize blood loss, operative time, and hepatic warm ischemia time. METHODS: Beginning in 1998 many major hepatic resections for hepatic tumors were performed with ultrasound-directed transparenchymal application of vascular linear cutting staplers. An endoscopic flexible neck vascular linear cutting stapler was used for control of the hepatic veins. RESULTS: From December 1998 to April 2003, 346 patients undergoing hepatic resection using this technique were identified from a prospective hepatobiliary tumor surgery database. Records were reviewed for blood loss, transfusion requirement, inflow occlusion (Pringle maneuver) time, overall operative time, and perioperative and postoperative complications. The average blood loss for all patients was 396 +/- 28.4 mL. The inflow occlusion time was 13.7 +/- .64 minutes with a total operative time of 140.7 +/- 3.7 minutes. Additional liver-related procedures were performed in 52% of the patients. The overall complication rate was 29.5% with a 90-day mortality rate of 1.4%. CONCLUSIONS: Ultrasound-directed transparenchymal application of vascular staplers to control inflow and outflow during major liver resection minimizes blood loss, warm ischemia time, and operative time compared to published reports of patients undergoing resection using other techniques.

Blood Loss, Surgical↗
PubMed2004

Modifications of human betaA1/betaA3-crystallins include S-methylation, glutathiolation, and truncation.

Disulfide bonding of lens crystallins contributes to the aggregation and insolubilization of these proteins that leads to cataract. A high concentration of reduced glutathione is believed to be key in preventing oxidation of crystallin sulfhydryls to form disulfide bonds. This protective role is decreased in aged lenses because of lower glutathione levels, especially in the nucleus. We recently found that human gamma-crystallins undergo S-methylation at exposed cysteine residues, a reaction that may prevent disulfide bonding. We report here that betaA1/A3-crystallins are also methylated at specific cysteine residues and are the most heavily methylated of the human lens crystallins. Among the methylated sites, Cys 64, Cys 99, and Cys 167 of betaA1-crystallin, methylation at Cys 99 is highest. Cys 64 and Cys 99 are also glutathiolated, even in a newborn lens. These post-translational modifications of the exposed cysteines may be important for maintaining the crystallin structure required for lens transparency. Previously unreported N-terminal truncations were also found.

Adult↗
PubMed2004

The risk of a mosquito-borne infection in a heterogeneous environment.

A common assumption about malaria, dengue, and other mosquito-borne infections is that the two main components of the risk of human infection--the rate at which people are bitten (human biting rate) and the proportion of mosquitoes that are infectious--are positively correlated. In fact, these two risk factors are generated by different processes and may be negatively correlated across space and time in heterogeneous environments. Uneven distribution of blood-meal hosts and larval habitat creates a spatial mosaic of demograPhic sources and sinks. Moreover, mosquito populations fluctuate temporally, forced by environmental variables such as rainfall, temperature, and humidity. These sources of spatial and temporal heterogeneity in the distribution of mosquito populations generate variability in the human biting rate, in the proportion of mosquitoes that are infectious, and in the risk of human infection. To understand how heterogeneity affects the epidemiology of mosquito-borne infections, we developed a set of simple models that incorporate heterogeneity in a stepwise fashion. These models predict that the human biting rate is highest shortly after the mosquito densities peak, near breeding sites where adult mosquitoes emerge, and around the edges of areas where humans are aggregated. In contrast, the proportion of mosquitoes that are infectious reflects the age structure of mosquito populations; it peaks where old mosquitoes are found, far from mosquito breeding habitat, and when mosquito population density is declining. Finally, we show that estimates for the average risk of infection that are based on the average entomological inoculation rate are strongly biased in heterogeneous environments.

Animals↗
PubMed2004

Multi-state unfolding of the alpha subunit of tryptophan synthase, a TIM barrel protein: insights into the secondary structure of the stable equilibrium intermediates by hydrogen exchange mass spectrometry.

The urea-induced unfolding of the alpha subunit of tryptophan synthase (alphaTS) from Escherichia coli, an eight-stranded (beta/alpha)(8) TIM barrel protein, has been shown to involve two stable equilibrium intermediates, I1 and I2, well populated at approximately 3 M and 5 M urea, respectively. The characterization of the I1 intermediate by circular dichroism (CD) spectroscopy has shown that I1 retains a significant fraction of the native ellipticity; the far-UV CD signal for the I2 species closely resembles that of the fully unfolded form. To obtain detailed insight into the disruption of secondary structure in the urea-induced unfolding process, a hydrogen exchange-mass spectrometry study was performed on alphaTS. The full-length protein was destabilized in increasing concentration of urea, the amide hydrogen atoms were pulse-labeled with deuterium, the labeled samples were quenched in acid and the products were analyzed by electrospray ionization mass spectrometry. Consistent with the CD results, the I1 intermediate protects up to approximately 129 amide hydrogen atoms against exchange while the I2 intermediate offers no protection. Electrospray ionization mass spectrometry analysis of the peptic fragments derived from alphaTS labeled at 3 M urea indicates that most of the region between residues 12-130, which constitutes the first four beta strands and three alpha helices, (beta/alpha)(1-3)beta(4), is structured. The (beta/alpha)(1-3)beta(4) module appears to represent the minimum sub-core of stability of the I1 intermediate. A 4+2+2 folding model is proposed as a likely alternative to the earlier 6+2 folding mechanism for alphaTS.

Escherichia coli↗
PubMed2004

Statics and dynamics of malaria infection in Anopheles mosquitoes.

The classic formulae in malaria epidemiology are reviewed that relate entomological parameters to malaria transmission, including mosquito survivorship and age-at-infection, the stability index (S), the human blood index (HBI), proportion of infected mosquitoes, the sporozoite rate, the entomological inoculation rate (EIR), vectorial capacity (C) and the basic reproductive number (R0). The synthesis emphasizes the relationships among classic formulae and reformulates a simple dynamic model for the proportion of infected humans. The classic formulae are related to formulae from cyclical feeding models, and some inconsistencies are noted. The classic formulae are used to to illustrate how malaria control reduces malaria transmission and show that increased mosquito mortality has an effect even larger than was proposed by Macdonald in the 1950's.

Animals↗
PubMed2004

Conformational dynamics of the GdmHCl-induced molten globule state of creatine kinase monitored by hydrogen exchange and mass spectrometry.

Our understanding of the mechanism of protein folding can be improved by the characterization of folding intermediate states. Intrinsic tryptophan fluorescence measurements of equilibrium GdmHCl-induced unfolding of MM-CK allow for the construction of a "phase diagram", which shows the presence of five different conformational states, including three partially folded intermediates. However, only three states are detected by using pulsed-labeled H-D exchange analyzed by electrospray ionization mass spectrometry. One of them is the native state, and the two other species are present in proportions strongly dependent on the GdmHCl concentration and denaturation time. The low-mass peak is due to a largely exchange-incompetent state, which has gained only approximately 10 deuteriums more than the native protein. This population of MM-CK molecules has undergone a small conformational change induced by low GdmHCl concentrations. However, this limited change is in itself not sufficient to inactivate the enzyme or is easily reversible. The high-mass peak corresponds to a population of MM-CK that is fully deuterated. The comparison of fluorescence, activity, and H-D exchange measurements shows that the maximally populated intermediate at 0.8 M GdmHCl has the characteristics of a molten globule. It has no activity; it has 55% of its native alpha-helices and a maximum fluorescence emission wavelength of approximately 341 nm, and it binds ANS strongly. However, no protection against exchange is detected under the conditions used in this work. This paradox, the presence of significant residual secondary and tertiary structures detected by optical probes and the total deuteration of its amide protons detected by H-D exchange and mass spectrometry, could be explained by a highly dynamic MM-CK molten globule.

Animals↗
PubMed2004

Projected benefits of active surveillance for vancomycin-resistant enterococci in intensive care units.

Hospitals use many strategies to control nosocomial transmission of vancomycin-resistant enterococci (VRE). Strategies include "passive surveillance," with isolation of patients with known previous or current VRE colonization or infection, and "active surveillance," which uses admission cultures, with subsequent isolation of patients who are found to be colonized with VRE. We created a mathematical model of VRE transmission in an intensive care unit (ICU) using data from an existing active surveillance program; we used the model to generate the estimated benefits associated with active surveillance. Simulations predicted that active surveillance in a 10-bed ICU would result in a 39% reduction in the annual incidence of VRE colonization when compared with no surveillance. Initial isolation of all patients, with withdrawal of isolation if the results of surveillance cultures are negative, was predicted to result in a 65% reduction. Passive surveillance was minimally effective. Using the best available data, active surveillance is projected to be effective for reducing VRE transmission in ICU settings.

Anti-Bacterial Agents↗
PubMed2004

Equilibrium and kinetic folding of rabbit muscle triosephosphate isomerase by hydrogen exchange mass spectrometry.

Unfolding and refolding of rabbit muscle triosephosphate isomerase (TIM), a model for (betaalpha)8-barrel proteins, has been studied by amide hydrogen exchange/mass spectrometry. Unfolding was studied by destabilizing the protein in guanidine hydrochloride (GdHCl) or urea, pulse-labeling with 2H2O and analyzing the intact protein by HPLC electrospray ionization mass spectrometry. Bimodal isotope patterns were found in the mass spectra of the labeled protein, indicating two-state unfolding behavior. Refolding experiments were performed by diluting solutions of TIM unfolded in GdHCl or urea and pulse-labeling with 2H2O at different times. Mass spectra of the intact protein labeled after one to two minutes had three envelopes of isotope peaks, indicating population of an intermediate. Kinetic modeling indicates that the stability of the folding intermediate in water is only 1.5 kcal/mol. Failure to detect the intermediate in the unfolding experiments was attributed to its low stability and the high concentrations of denaturant required for unfolding experiments. The folding status of each segment of the polypeptide backbone was determined from the deuterium levels found in peptic fragments of the labeled protein. Analysis of these spectra showed that the C-terminal half folds to form the intermediate, which then forms native TIM with folding of the N-terminal half. These results show that TIM folding fits the (4+4) model for folding of (betaalpha)8-barrel proteins. Results of a double-jump experiment indicate that proline isomerization does not contribute to the rate-limiting step in the folding of TIM.

Animals↗
PubMed2004

Persistent colonization and the spread of antibiotic resistance in nosocomial pathogens: resistance is a regional problem.

Infections with antibiotic-resistant bacteria (ARB) in hospitalized patients are becoming increasingly frequent despite extensive infection-control efforts. Infections with ARB are most common in the intensive care units of tertiary-care hospitals, but the underlying cause of the increases may be a steady increase in the number of asymptomatic carriers entering hospitals. Carriers may shed ARB for years but remain undetected, transmitting ARB to others as they move among hospitals, long-term care facilities, and the community. We apply structured population models to explore the dynamics of ARB, addressing the following questions: (i) What is the relationship between the proportion of carriers admitted to a hospital, transmission, and the risk of infection with ARB? (ii) How do frequently hospitalized patients contribute to epidemics of ARB? (iii) How do transmission in the community, long-term care facilities, and hospitals interact to determine the proportion of the population that is carrying ARB? We offer an explanation for why ARB epidemics have fast and slow phases and why resistance may continue to increase despite infection-control efforts. To successfully manage ARB at tertiary-care hospitals, regional coordination of infection control may be necessary, including tracking asymptomatic carriers through health-care systems.

Bacterial Infections↗
PubMed2004

A priori prediction of disease invasion dynamics in a novel environment.

Directly transmitted infectious diseases spread through wildlife populations as travelling waves away from the sites of original introduction. These waves often become distorted through their interaction with environmental and population heterogeneities and by long-distance translocation of infected individuals. Accurate a priori predictions of travelling waves of infection depend upon understanding and quantifying these distorting factors. We assess the effects of anisotropies arising from the orientation of rivers in relation to the direction of disease-front propagation and the damming effect of mountains on disease movement in natural populations. The model successfully predicts the local and large-scale prevaccination spread of raccoon rabies through New York State, based on a previous spatially heterogeneous model of raccoon-rabies invasion across the state of Connecticut. Use of this model provides a rare example of a priori prediction of an epidemic invasion over a naturally heterogeneous landscape. Model predictions matched to data can also be used to evaluate the most likely points of disease introduction. These results have general implications for predicting future pathogen invasions and evaluating potential containment strategies.

Animals↗
PubMed2004

Human telomerase reverse transcriptase (hTERT) and Ki-67 are better predictors of survival than established clinical indicators in patients undergoing curative hepatic resection for colorectal metastases.

BACKGROUND: We evaluated hTERT and Ki-67 expression in patients who underwent curative resection of hepatic colorectal metastases to determine if these markers of cell proliferation correlated better with survival than an established scoring system that is based on clinical predictors. METHODS: Patients operated on between 1993 and 1997 whose survival time was known were analyzed. For each patient, the clinical prognostic score was derived on the basis of primary node status, disease-free interval, number of hepatic tumors, largest tumor, and carcinoembryonic antigen level, and tumor specimens were analyzed for Ki-67 and hTERT with use of standard immunohistochemical techniques. The immunohistochemical analysis was blinded to all patient characteristics. RESULTS: The study included 66 patients. Twenty-six survived less than 2 years after surgery, 19 survived 2-5 years, and 21 survived more than 5 years. Ki-67 positivity and hTERT positivity (labeling indexes greater than or equal to 50%) were observed in 24 patients and 23 patients, respectively. The clinical score did not predict survival, although there was a weak trend toward a lower score in patients with better survival. Both Ki-67 (P =.04) and hTERT (P =.0001) correlated better with survival than did the clinical score. CONCLUSIONS: In patients undergoing curative resection of hepatic colorectal metastases, hTERT and Ki-67 are better predictors of survival than is a score based on clinical features.

Colorectal Neoplasms↗
PubMed2004

Unexpected products from the reaction of the synthetic cross-linker 3,3'-dithiobis(sulfosuccinimidyl propionate), DTSSP with peptides.

Synthetic cross-linking reagents, such as 3,3'-dithiobis(sulfosuccinimidyl propionate), DTSSP, can react with sidechains of amino acids that are within close proximity. Identification of cross-linked residues provides insight into the folded structures of proteins. However, analysis of proteolytic digests of proteins cross-linked with commercially available DTSSP is difficult because many ions cannot be attributed to reported reactions of DTSSP. To better understand the reactivity of DTSSP, products from the reaction of DTSSP with several model peptides were analyzed by HPLC electrospray ionization mass spectrometry (ESIMS). Several products not previously reported were identified. Sources for these unexpected products were traced to reaction of DTSSP with contaminant ammonium ions in the buffer, to reaction of contaminants present in the commercial DTSSP reagent, and to reactivity of DTSSP with serine and tyrosine residues. In addition, the collision-induced-dissociation (CID) of peptides modified by DTSSP was investigated. These results showed that certain DTSSP-peptide adducts easily undergo in-source fragmentation to give additional unexpected ions. This study of the reactions of DTSSP with model peptides has revealed the major types of ions that are likely to be found in proteolytic digests of proteins cross-linked with DTSSP, thereby facilitating identification of the cross-linked residues that can provide information about the three-dimensional structures of folded proteins.

Amines↗
PubMed2004

Coupling ecology and evolution: malaria and the S-gene across time scales.

Malaria has long been a scourge to humans. The exceptionally high mortality in some regions has led to strong selection for resistance, even at the cost of increased risk of potentially fatal red blood cell deformities in some offspring. In particular, genes that confers resistance to malaria when they appear in heterozygous individuals are known to lead to sickle-cell anemia, or other blood diseases, when they appear in homozygous form. Thus, there is balancing selection against the evolution of resistance, with the strength of that selection dependent upon malaria prevalence. Over longer time scales, the increased frequency of resistance in a population might be expected to decrease the frequency of malaria and reduce selection for resistance. However, possession of the sickle-cell gene leads to longer-lasting parasitaemia in heterozygote individuals, and therefore the presence of resistance may actually increase infection prevalence. In this paper, we explore the interplay among these processes, operating over very different time scales. In particular, we show that on the fast time scale of malarial dynamics, the disease level reaches an equilibrium; on the slower, evolutionary time scale, this equilibrium tracks gene frequency. We analyze the slow time scale dynamics to investigate the impact of malaria on the evolution of resistance.

Algorithms↗
PubMed2004

In vitro characterization of aortic retrograde and antegrade flow from pulsatile and non-pulsatile ventricular assist devices.

BACKGROUND: Many advances have been made in left ventricular assist device (LVAD) development including the introduction of smaller, non-pulsatile pumps. However, controversy exists over the potential significance of non-pulsatile blood flow. In addition, some newer LVADs incorporate descending aortic anastomosis (and therefore retrograde ascending aortic flow) for outflow rather than the traditional ascending aortic anastomosis. This, combined with non-pulsatile flow, may significantly increase the risks of ascending aortic thrombus formation, especially if native cardiac function is negligible and the aortic valve remains closed. The purpose of this study was to compare pulsatile and non-pulsatile flow generated by LVADs with outflow to the ascending aorta and descending aorta. METHODS: An in vitro mock circulatory loop, driven by either a pulsatile or a non-pulsatile LVAD, was anastomosed to transparent aortic models at either the ascending or descending aortic position. The aortic valve was kept closed, modeling no native cardiac output. Normal saline was used as a blood analog. Methylene blue dye was injected into the ascending aorta and aortic arch to demonstrate flow patterns. Dye washout time (in seconds) was used as a marker of flow stagnation and potential thrombogenicity. LVAD flow, rate, after-load and coronary flow were measured. RESULTS: Dye washout times at a flow rate of 5 liters/min were 1.7 +/- 0.75, 2.1 +/- 0.71, 4.7 +/- 0.82 and 9.9 +/- 4.4 seconds for pulsatile ascending (PA), non-pulsatile ascending (NPA), pulsatile descending (PD) and non-pulsatile descending flow (NPD), respectively. Coronary flow averaged 294 ml/min over all set-ups. Dye washout times at a flow rate of 4-liters/min were 3.0 +/- 1.0, 3.0 +/- 0.8, 14.0 +/- 3.8 and 25.0 +/- 9.1 seconds for PA, NPA, PD and NPD, respectively. Coronary flow averaged 227 ml/min over all set-ups. Ascending aortic anastomoses were associated with shorter dye washout times compared with descending aortic anastomoses, regardless of flow type (p < 0.001). There was no difference in washout time between pulsatile and non-pulsatile flow in the ascending aortic position (p = 0.23 and 0.12 for 5 and 4 liters/min, respectively). Pulsatile flow in the descending aorta had shorter washout times than non-pulsatile flow in the descending aorta (p < 0.001 and p = 0.004 for 5 and 4 liters/min, respectively). CONCLUSIONS: LVAD descending aortic anastomosis and retrograde aortic flow is associated with increased flow stagnation in the ascending aorta. This may increase the risk for thrombus formation in patients relying solely on retrograde aortic flow, especially if cardiac function and antegrade blood flow returns.

Aorta↗