Biomedical subjects

Smita Patel

Publications and source records attributed to Smita Patel.

At least 19 recordsLinked to original sources

PubMed2006

Autotomic behavior of the propeptide in propeptide-mediated folding of prosubtilisin E.

The 77 residue propeptide at the N-terminal end of subtilisin E plays an essential role in subtilisin folding as a tailor-made intramolecular chaperone. Upon completion of folding, the propeptide is autoprocessed and removed by subtilisin digestion. This propeptide-mediated protein folding has been used as a paradigm for the study of protein folding. Here, we show by three independent methods, that the propeptide domain and the subtilisin domain show distinctive intrinsic stability that is obligatory for efficient autoprocessing of the propeptide domain. Two tryptophan residues, Trp106 and Trp113, on the surface of subtilisin located on one of the two helices that form the interface between the propeptide and the subtilisin domains play a key role in maintaining the distinctive instability of the propeptide domain, after completion of folding. When either of the Trp residues was substituted with Tyr, the characteristic biphasic heat denaturation profile of two domains unfolding was not observed, resulting in a single transition of denaturation. The results provide evidence that the propeptide not only plays an essential role in subtilisin folding, but upon completion of folding it behaves as an independent domain. Once the propeptide-mediated folding is completed, the propeptide domain is readily eliminated without interference from the subtilisin domain. This "autotomic" behavior of the propeptide may be a prevailing principle in propeptide-mediated protein folding.

Circular Dichroism↗

PubMed2006

Mg2+ and a key lysine modulate exchange activity of eukaryotic translation elongation factor 1B alpha.

To sustain efficient translation, eukaryotic elongation factor B alpha (eEF1B alpha) functions as the guanine nucleotide exchange factor for eEF1A. Stopped-flow kinetics using 2'-(or 3')-O-N-methylanthraniloyl (mant)-GDP showed spontaneous release of nucleotide from eEF1A is extremely slow and accelerated 700-fold by eEF1B alpha. The eEF1B alpha-stimulated reaction was inhibited by Mg2+ with a K(1/2) of 3.8 mM. Previous structural studies predicted the Lys-205 residue of eEF1B alpha plays an important role in promoting nucleotide exchange by disrupting the Mg2+ binding site. Co-crystal structures of the lethal K205A mutant in the catalytic C terminus of eEF1B alpha with eEF1A and eEF1A.GDP established that the lethality was not due to a structural defect. Instead, the K205A mutant drastically reduced the nucleotide exchange activity even at very low concentrations of Mg2+. A K205R eEF1B alpha mutant on the other hand was functional in vivo and showed nearly wild-type nucleotide dissociation rates but almost no sensitivity to Mg2+. These results indicate the significant role of Mg2+ in the nucleotide exchange reaction by eEF1B alpha and establish the catalytic function of Lys-205 in displacing Mg2+ from its binding site.

Catalysis↗

PubMed2006

A new class of selective, non-basic 5-HT2A receptor antagonists.

Based on an existing series of 5-HT2A receptor ligands containing a basic nitrogen, we designed a non-basic lead that had reduced affinity for both the 5-HT2A receptor and the IKr potassium channel. The present paper describes the development of this lead to a novel series of non-basic piperidine sulfonamides and amides that have high affinity for the 5-HT2A receptor, whilst maintaining excellent selectivity over off target activities such as the IKr channel. This work has shown that the proposed pharmacophore model for the 5-HT2A receptor which suggests that a basic nitrogen is required for the binding of ligands is questionable.

Humans↗

PubMed2006

Multidetector row CT pulmonary angiography and indirect venography for the diagnosis of venous thromboembolic disease in intensive care unit patients.

RATIONALE AND OBJECTIVES: To determine the diagnostic quality, performance characteristics and interreader reliability of computed tomography pulmonary angiography (CTPA) and venography (CTV) in intensive care unit (ICU) patients with suspected venous thromboembolism (VTE). MATERIALS AND METHODS: A total of 100 consecutive ICU patient CT examinations performed for clinically suspected VTE on a four-row CT scanner were reviewed. Three readers rated the diagnostic quality of each CTPA and CTV examination as excellent, acceptable, or nondiagnostic. Readers scored the overall determination for pulmonary embolism (PE) and deep venous thrombosis (DVT) using a 5-point scale, and scored the determination for PE by anatomic level. Receiver operator characteristic (ROC) analysis was performed for each reader and the original clinical report, using consensus interpretation as the reference standard. Interobserver variability for PE and DVT was determined using kappa analysis, and was stratified by examination quality. RESULTS: A total of 25% of CTPA examinations were nondiagnostic, most commonly because of motion artifact and poor contrast opacification. A total of 24% of CTV examinations were nondiagnostic, most commonly because of poor contrast opacification and metallic hardware. Using receiver operating characteristic analysis, the areas under the curve (Az) for PE diagnosis were 0.875, 0.923, 0.888, and 0.674 for the three readers and clinical reading, respectively, and for DVT diagnosis were 0.842, 0.859, 0.952 and 0.669. Interobserver agreement for detection of PE was moderate at the supralobar level (kappa = 0.55), very good at the lobar level (kappa = 0.69), and moderate for segmental (kappa = 0.54) and subsegmental arteries (kappa = 0.44). Overall reader agreement was good for excellent/good quality CTPA examinations (kappa = 0.52-0.56), and poor when examination quality was poor (kappa = 0.06). CONCLUSIONS: CTPA and CTV are sufficiently accurate and reliable techniques for evaluating VTE in ICU patients, particularly in light of patient complexity.

Adolescent↗

PubMed2005

A negative carotid plaque area test is superior to other noninvasive atherosclerosis studies for reducing the likelihood of having underlying significant coronary artery disease.

OBJECTIVE: Coronary calcium score (CCS), carotid plaque area (CPA), intima-media thickness (IMT), and C-reactive protein (CRP) are independent predictors of cardiovascular prognosis. Although each test may enhance risk stratification, their comparative abilities to screen for underlying coronary stenoses in individual patients is less established. METHODS AND RESULTS: Forty-two patients who had a 16-slice coronary computed tomography angiogram (CTA) performed were invited to have CPA, IMT, and CRP measured. CPA was defined as the sum of all the cross-sectional areas of each plaque >1 mm in diameter found in all carotid vessels bilaterally. CCS and the number plus degree of stenotic coronary arteries were determined by CTA. The presence of clinically significant coronary artery disease (CAD) was defined as the existence of any stenosis > or =50%. CTA identified clinically significant CAD in 43% of the patients. CPA >0 was more sensitive (72%) and specific (58%) than a CCS >0 (58% and 55%) for identifying CAD. A "clean" carotid artery (CPA=0) provides a superior negative predictive value (74%) and likelihood ratio of a negative test (0.48) than all other studies, in particular versus a CCS=0 (65% and 0.72). The areas under the receiver-operator curves for CPA and CCS in relation to any CAD were similar (0.640 versus 0.675). Carotid IMT and CRP performed poorly compared with CPA and CCS. For detecting CAD in only the left main or left anterior descending artery, the negative predictive value and likelihood ratio of a negative test remained superior for CPA (87% and 0.33) compared with CCS (80% and 0.56). In our population with a prevalence of these coronary lesions of 30%, the post-test probability in any patient with a negative CPA result is reduced to 10%. CONCLUSIONS: CPA determination is superior to CCS, IMT, and CRP in its ability to reduce the likelihood of clinically significant underlying CAD in patients of varying cardiac risk.

Aged↗

PubMed2005

Lack of direct role of parkin in the steady-state level and aggregation of alpha-synuclein and the clearance of pre-formed aggregates.

Mutations in parkin and alpha-synuclein (alpha-syn) are linked to heritable forms of Parkinson's disease (PD). Recently, it has been shown that parkin mitigates alpha-syn-induced neuronal cell death in animal and tissue culture models, suggesting that there is a functional relationship between these two proteins. Although the mechanism by which parkin protects cells from alpha-syn-induced cytotoxicity remains elusive, it is tempting to speculate that parkin might directly regulate the normal metabolism and aggregation of alpha-syn. In the current study, we show that neither the suppression of endogenous parkin expression nor ectopic overexpression affects the steady-state levels of endogenous alpha-syn expression, overall aggregation of this protein, or breakdown of pre-formed aggregates in human neuroblastoma cells. These results suggest that parkin is not directly involved in the metabolism of alpha-syn, its aggregation, or the clearance of pre-formed aggregates.

Analysis of Variance↗

PubMed2005

4-Fluorosulfonylpiperidines: selective 5-HT2A ligands for the treatment of insomnia.

Incorporation of fluorine at the 4-position of an existing series of sulfonyl piperidine 5-HT2A antagonists gave compounds with increased selectivity over the IKr potassium channel. This work led to the identification of 3b, a compound that gave no increase in QTc in the anesthetized dog up to plasma levels as high as 148 microM. Furthermore, 3b has been shown to increase slow-wave sleep bout duration and to decrease the number of awakenings in rats, indicating the potential utility of 5-HT2A antagonists in the treatment of insomnia.

Animals↗

PubMed2005

Intravesicular localization and exocytosis of alpha-synuclein and its aggregates.

Alpha-synuclein (alpha-syn), particularly in its aggregated forms, is implicated in the pathogenesis of Parkinson's disease and other related neurological disorders. However, the normal biology of alpha-syn and how it relates to the aggregation of the protein are not clearly understood. Because of the lack of the signal sequence and its predominant localization in the cytosol, alpha-syn is generally considered exclusively an intracellular protein. Contrary to this assumption, here, we show that a small percentage of newly synthesized alpha-syn is rapidly secreted from cells via unconventional, endoplasmic reticulum/Golgi-independent exocytosis. Consistent with this finding, we also demonstrate that a portion of cellular alpha-syn is present in the lumen of vesicles. Importantly, the intravesicular alpha-syn is more prone to aggregation than the cytosolic protein, and aggregated forms of alpha-syn are also secreted from cells. Furthermore, secretion of both monomeric and aggregated alpha-syn is elevated in response to proteasomal and mitochondrial dysfunction, cellular defects that are associated with Parkinson's pathogenesis. Thus, intravesicular localization and secretion are part of normal life cycle of alpha-syn and might also contribute to pathological function of this protein.

Animals↗

PubMed2005

Distribution of brain atrophy in behavioral variant frontotemporal dementia.

Marked brain atrophy occurs in frontotemporal dementia (FTD) yet substantial variation between cases is seen. Recently, a four-level staging scheme which reflects increasing disease duration, severity of dementia and degree of neurodegeneration was described. In the present study, the extent and magnitude of atrophy in behavioral variant FTD and its relationship to disease duration and pathological subtype was further evaluated by quantifying the volume of 30 anatomically-defined regions. A validated point count technique was applied to 17 patients with FTD (9 Pick's disease, 6 dementia lacking distinctive histology, 2 FTD with motor neuron disease) and 21 controls. Atrophy was seen in all brain regions except the inferior frontal cortex and area 37. As might be expected, increasing severity of atrophy occurred with increasing disease duration and stage however measurable atrophy was more widespread than indicated by the staging scheme. Furthermore, severity of atrophy was not related to pathological subtype. Frontal, limbic and temporal regions appeared to be severely affected early in the disease process with temporal lobe atrophy the best predictor of disease duration. White matter, more posterior regions and the subcortex were affected later in the disease. These findings demonstrate a pattern of selective vulnerability which progresses over time. Furthermore, they demonstrate that although patients with a similar clinical subtype may have differing underlying histopathology, the pattern, severity and progression of brain atrophy is the same. This suggests that the regional pattern of neurodegeneration, rather than the type of histopathology influences the clinical syndrome in FTD.

Aged↗

PubMed2005

Estimating the effects of informal radiology resident teaching on radiologist productivity: what is the cost of teaching?

RATIONALE AND OBJECTIVES: One mission of an academic radiology department is to teach. The greatest teaching effort is directed at radiology residents. As clinical work demands increase, informal, non-revenue-generating, teaching may suffer. We sought to determine the economic consequences of teaching. MATERIALS AND METHODS: With the use of a picture archiving and communications system, 6 radiology faculty members independently interpreted and dictated digitally acquired bone and chest radiographs for 1 hour alone and again 10-12 weeks later with a first-year resident. During the second session, the quality of teaching was graded by independent observers. The number of cases, relative value units (RVUs), and reimbursement for each session were calculated. RESULTS: The difference in number of cases dictated working alone (mean, 44.7) and with a first-year resident (mean, 23.5) was significant (P = 0.007). The difference between RVUs generated by faculty alone (mean, 9.0) and with a resident (mean, 4.5) also was significant (P = 0.006), and the difference in dollars billed when working alone (mean, $1558.45) and with a resident (mean, $777.65) was significant (P = 0.007). As teaching quality increased, the number of cases interpreted, dollars billed, and RVUs trended lower. CONCLUSION: Informal resident teaching significantly reduces clinical throughput, reducing examination volume, RVUs, and dollars billed by approximately half.

Bone and Bones↗

PubMed2005

Preliminary investigation of computer-aided detection of pulmonary embolism in three-dimensional computed tomography pulmonary angiography images.

RATIONALE AND OBJECTIVES: We sought to develop a computer-aided diagnosis (CAD) system for assisting radiologists in the detection of pulmonary embolism (PE) on computed tomography pulmonary angiographic (CTPA) images. MATERIALS AND METHODS: An adaptive three-dimensional (3D) voxel clustering method was developed based on expectation-maximization (EM) analysis to extract vessels from their surrounding tissues. Using a connected component analysis, the vessel tree was reconstructed by tracking the vessel and its branches in 3D space. The tracked vessels were prescreened for suspicious PE areas using a second EM analysis. A rule-based false-positive (FP) reduction method was designed to detect true PE based on the features of PE and vessels. In this preliminary study, 14 patients with positive CTPA for PE were studied. CT scans were performed at 1.25-mm collimation using a GE LightSpeed CT scanner; eight of these patients also had extensive lung parenchymal or pleural disease. One hundred sixty-three emboli were identified by two experienced thoracic radiologists. The emboli identified by the radiologists were used as the "gold standard." For each embolus, the percent diameter occlusion (clot) and conspicuity of embolus (rating of 1 to 5, with 5 being the most conspicuous) were visually estimated. One hundred one emboli were identified in the six patients without lung diseases; 57 were proximal to the subsegmental and 44 were subsegmental. For the eight patients with lung diseases, 62 emboli were identified, of which 37 were proximal to the subsegmental and 25 were subsegmental. A computer-detected volume was counted as true-positive when it overlapped with an embolus volume identified by the radiologists. RESULTS: In the cases without lung diseases, if the PE had a conspicuity of >2 and only partially (20%-80%) occluded the vessel, our method detected 92.0% of proximal emboli and 77.8% of subsegmental emboli, with an average of 18.3 FPs/case. In the cases containing extensive lung disease, 66.7% and 40.0% of the PEs were detected with an average of 11.4 FPs/case under the same conditions. For the 14 PE cases, 13 of them were diagnosed as positive PE cases (case sensitivity was 92.9%). CONCLUSION: This preliminary study indicates that our automated method is a promising approach to CAD of PE on CTPA. Further study is under way to collect a larger data set and to improve the detection accuracy for PE, especially those with <20% or >80% occlusion, and for very subtle PE. A fully developed CAD system is expected to provide a useful aid for PE detection on CTPA.

Angiography↗

PubMed2005

Human immunodeficiencies that predispose to intracellular bacterial infections.

PURPOSE OF REVIEW: Patients treated with anti-tumour necrosis factor agents have an increased risk of active tuberculosis. Mycobacteria are bacterial pathogens capable of surviving and multiplying within macrophages; these infections are characterised by granulomatous inflammation. This review addresses the effects of inherited and acquired immunodeficiencies on the susceptibility to the development of intracellular bacterial infections. RECENT FINDINGS: Primary and secondary immunodeficiencies that result in severely impaired T cell function or macrophage activation result in an increased risk of mycobacterial and Salmonella infection. Conversely, inherited or acquired antibody or complement deficiency does not lead to increased susceptibility to these pathogens. Inherited defects in the interleukin-12/interleukin-23-dependent interferon-gamma pathway due to mutations in genes encoding the p40 chain common to interleukin-12 and interleukin-23, the beta1 chain shared by interleukin-12 and interleukin-23 receptors, interferon-gamma receptor chains 1 or 2, or signal transducer and activator of transcription, predispose to severe infections caused by poorly pathogenic mycobacteria and Salmonella species. Acquired defects of cytokine function causing increased susceptibility to these pathogens include anti-tumor necrosis factor therapy and the generation of interferon-gamma-neutralising autoantibodies. Defective nuclear factor kappaB activation caused by hypomorphic mutations of the nuclear factor kappaB essential modulator gene, which compromises the function of Toll receptors, interleukin-IL receptors, and tumor necrosis factor-alpha receptors, also increases susceptibility to severe mycobacterial infections. Patients with inherited defects in the phagocyte nicotine-adenine dinucleotide phosphate oxidase system are highly susceptible to Salmonella infections but only exhibit slightly increased susceptibility to mycobacteria. SUMMARY: Collectively, these observations highlight immune mechanisms that are essential for protection against intracellular bacteria. This information provides clinicians with a framework for investigating patients with potentially life-threatening intracellular bacterial infections.

Disease Susceptibility↗

PubMed2005

Helical CT for the evaluation of acute pulmonary embolism.

OBJECTIVE: In this article, we review the current role of CT pulmonary angiography and indirect CT venography for the evaluation of pulmonary thromboembolic disease. CONCLUSION: With advances in MDCT technology, evaluation of pulmonary thromboembolic disease can now be performed with combined CT pulmonary angiography and CT venography as a "one-stop-shopping" test. CT pulmonary angiography is cost-effective, is accurate, has high interobserver agreement, and has an added advantage of detecting other life-threatening diseases in the chest that mimic pulmonary embolism.

Acute Disease↗

PubMed2004

Insights into the role of an active site aspartate in Ty1 reverse transcriptase polymerization.

Long terminal repeat-containing retrotransposons encode reverse transcriptases (RTs) that replicate their RNA into integratable, double-stranded DNA. A mutant version of the RT from Saccharomyces cerevisiae retrotransposon Ty1, in which one of the three active site aspartates has been changed to asparagine (D211N), is still capable of in vitro polymerization, although it is blocked for in vivo transposition. We generated recombinant WT and D211N Ty1 RTs to study RT function and determine specific roles for the Asp(211) residue. Presteady-state kinetic analysis of the two enzymes shows that the D211N mutation has minimal effect on nucleotide binding but reduces the k(pol) by approximately 230-fold. The mutation reduces binding affinity for both Mn(2+) and Mg(2+), indicating that the Asp(211) side chain helps create a tight metal binding pocket. Although both enzymes are highly processive and tend to remain bound to their initial substrate, each shows distinctive patterns of pausing, attributable to interactions between metal ions and the active site residue. These results provide insights to specific roles for the Asp(211) residue during polymerization and indicate unusual enzymatic properties that bear on the Ty1 replication pathway.

Aspartic Acid↗

PubMed2004

Survival advantage of cultured human vascular endothelial cells that lost chromosome 13.

We explored the nature of chromosome 13 loss in cultured human vascular endothelial cells (HUVECs). Chromosome 13 loss detected by metaphase and interphase analysis was noted in earlier passages of HUVEC strains with no relation to telomere length or replicative senescence. Ectopic expression of telomerase did not influence the loss of chromosome 13. HUVECs losing chromosome 13 demonstrated increased migratory potential and loss of heterozygosity. Collectively, these observations suggest that the loss of chromosome 13 gives cultured HUVECs a replicative advantage.

Cell Count↗

PubMed2004

Clearance of alpha-synuclein oligomeric intermediates via the lysosomal degradation pathway.

Cytoplasmic deposition of alpha-synuclein aggregates is a common pathological feature of many neurodegenerative diseases. Strong evidence for the causative role of alpha-synuclein in these disorders is provided by genetic linkage between this gene and familial Parkinson's disease and by neurodegeneration in transgenic animals that overexpress this protein. In particular, it has been hypothesized that the accumulation of nonfibrillar oligomers of alpha-synuclein, which serve as intermediates for fibrillar inclusion body formation, causes neurodegeneration. However, little is known about how cells handle potentially toxic protein aggregates. Here we demonstrate that cells are capable of clearing preformed alpha-synuclein aggregates via the lysosomal degradation pathway. Consequently, blocking this pathway causes the accumulation of the aggregates in non-neuronal cells, differentiated neuroblastoma cells, and primary cortical neurons. This aggregate clearance occurs in an aggregation stage-specific manner; oligomeric intermediates are susceptible to clearance, whereas mature fibrillar inclusion bodies are not. Neutralization of the acidic compartments leads to the accumulation of alpha-synuclein aggregates and exacerbates alpha-synuclein toxicity in postmitotic neuronal cells, suggesting that the accumulation of oligomeric intermediates may be an important event leading to alpha-synuclein-mediated cell death. These results suggest that enhancing lysosomal function may be a potential therapeutic strategy to halt or even prevent the pathogenesis of Parkinson's disease and other Lewy body diseases.

Animals↗

PubMed2004

Antisense RNA regulation by stable complex formation in the Enterococcus faecalis plasmid pAD1 par addiction system.

The par stability determinant, encoded by the Enterococcus faecalis plasmid pAD1, is the only antisense RNA regulated postsegregational killing system identified in gram-positive bacteria. Because of the unique organization of the par locus, the par antisense RNA, RNA II, binds to its target, RNA I, at relatively small, interspersed regions of complementarity. The results of this study suggest that, rather than targeting the antisense bound message for rapid degradation, as occurs in most other antisense RNA regulated systems, RNA I and RNA II form a relatively stable, presumably translationally inactive complex. The stability of the RNA I-RNA II complex would allow RNA I to persist in an untranslated state unless or until the encoding plasmid was lost. After plasmid loss, RNA II would be removed from the complex, allowing translational activation of RNA I. The mechanism of RNA I activation in vivo is unknown, but in vitro dissociation experiments suggest that active removal of RNA II, for example by a cellular RNase, may be required.

Amino Acid Sequence↗

PubMed2004

Gender differences in hip anatomy: possible implications for injury tolerance in frontal collisions.

Male occupants in frontal motor vehicle collisions have reduced tolerance for hip fractures than females in similar crashes. We studied 92 adult pelvic CT scans and found significant gender differences in bony pelvic geometry, including acetabular socket depth and femoral head width. Significant differences were also noted in the presentation angle of the acetabular socket to frontal loading. The observed differences provide biomechanical insight into why hip injury tolerance may differ with gender. These findings have implications for the future design of vehicle countermeasures as well as finite element models capable of more accurately predicting body tolerances to injury.

Accidents, Traffic↗