bluuebunny/crossref_metadata_2025 huggingface dataset:
STING Millennium Suite: integrated software for extensive analyses of 3d structures of proteins and their complexes
Authors: Higa Roberto H , Togawa Roberto C , Montagner Arnaldo J , Palandrani Juliana CF , Okimoto Igor KS , Kuser Paula R , Yamagishi Michel EB , Mancini Adauto L , Neshich Goran
Published: August 2004

Background

The integration of many aspects of protein/DNA structure analysis is an important requirement for software products in general area of structural bioinformatics. In fact, there are too few software packages on the internet which can be described as successful in this respect. We might say that what is still missing is publicly available, web based software for interactive analysis of the sequence/structure/function of proteins and their complexes with DNA and ligands. Some of existing software packages do have certain level of integration and do offer analysis of several structure related parameters, however not to the extent generally demanded by a user.

Results

We are reporting here about new Sting Millennium Suite (SMS) version which is fully accessible (including for local files at client end), web based software for molecular structure and sequence/structure/function analysis. The new SMS client version is now operational also on Linux boxes and it works with non-public pdb formatted files (structures not deposited at the RCSB/PDB), eliminating earlier requirement for the registration if SMS components were to be used with user's local files. At the same time the new SMS offers some important additions and improvements such as link to ProTherm as well as significant re-engineering of SMS component ConSSeq. Also, we have added 3 new SMS mirror sites to existing network of global SMS servers: Argentina, Japan and Spain.

Conclusion

SMS is already established software package and many key data base and software servers worldwide, do offer either a link to, or host the SMS. SMS (S ting M illennium S uite) is web-based publicly available software developed to aid researches in their quest for translating information about the structures of macromolecules into knowledge. SMS allows to a user to interactively analyze molecular structures, cross-referencing visualized information with a correlated one, available across the internet. SMS is already used as a didactic tool by some universities. SMS analysis is now possible on Linux OS boxes and with no requirement for registration when using local files.
Extrinsic factors regulate partial agonist efficacy of strychnine-sensitive glycine receptors
Authors: Farroni Jeffrey S , McCool Brian A
Published: August 2004

Background

Strychnine-sensitive glycine receptors in many adult forebrain regions consist of alpha2 + beta heteromeric channels. This subunit composition is distinct from the alpha1 + beta channels found throughout the adult spinal cord. Unfortunately, the pharmacology of forebrain alpha2beta receptors are poorly defined compared to 'neonatal' alpha2 homomeric channels or 'spinal' alpha1beta heteromers. In addition, the pharmacologic properties of native alpha2beta glycine receptors have been generally distinct from receptors produced by heterologous expression. To identify subtype-specific pharmacologic tools for the forebrain alpha2beta receptors, it is important to identify a heterologous expression system that closely resembles these native glycine-gated chloride channels.

Results

While exploring pharmacological properties of alpha2beta glycine receptors compared to alpha2-homomers, we found that distinct heterologous expression systems appeared to differentially influence partial agonist pharmacology. The β-amino acid taurine possessed 30–50% efficacy for alpha2-containing receptor isoforms when expressed in HEK 293 cells. However, taurine efficacy was dramatically reduced in L-cell fibroblasts. Similar results were obtained for β-alanine. The efficacy of these partial agonists was also strongly reduced by the beta subunit. There were no significant differences in apparent strychnine affinity values calculated from concentration-response data between expression systems or subunit combinations. Nor did relative levels of expression correlate with partial agonist efficacy when compared within or between several different expression systems. Finally, disruption of the tubulin cytoskeleton reduced the efficacy of partial agonists in a subunit-dependent, but system-independent, fashion.

Conclusions

Our results suggest that different heterologous expression systems can dramatically influence the agonist pharmacology of strychnine-sensitive glycine receptors. In the systems examine here, these effects are independent of both absolute expression level and any system-related alterations in the agonist binding site. We conclude that complex interactions between receptor composition and extrinsic factors may play a significant role in determining strychnine-sensitive glycine receptor partial agonist pharmacology.
DMBT1 expression is down-regulated in breast cancer
Authors: Braidotti P , Nuciforo PG , Mollenhauer J , Poustka A , Pellegrini C , Moro A , Bulfamante G , Coggi G , Bosari S , Pietra GG
Published: August 2004

Background

We studied the expression of DMBT1 (deleted in malignant brain tumor 1), a putative tumor suppressor gene, in normal, proliferative, and malignant breast epithelium and its possible relation to cell cycle.

Methods

Sections from 17 benign lesions and 55 carcinomas were immunostained with anti DMBT1 antibody (DMBTh12) and sections from 36 samples, were double-stained also with anti MCM5, one of the 6 pre-replicative complex proteins with cell proliferation-licensing functions. DMBT1 gene expression at mRNA level was assessed by RT-PCR in frozen tissues samples from 39 patients.

Results

Normal glands and hyperplastic epithelium in benign lesions displayed a luminal polarized DMBTh12 immunoreactivity. Normal and hyperplastic epithelium adjacent to carcinomas showed a loss of polarization, with immunostaining present in basal and perinuclear cytoplasmic compartments. DMBT1 protein expression was down-regulated in the cancerous lesions compared to the normal and/or hyperplastic epithelium adjacent to carcinomas (3/55 positive carcinomas versus 33/42 positive normal/hyperplastic epithelia; p = 0.0001). In 72% of cases RT-PCR confirmed immunohistochemical results. Most of normal and hyperplastic mammary cells positive with DMBTh12 were also MCM5-positive.

Conclusions

The redistribution and up-regulation of DMBT1 in normal and hyperplastic tissues flanking malignant tumours and its down-regulation in carcinomas suggests a potential role in breast cancer. Moreover, the concomitant expression of DMTB1 and MCM5 suggests its possible association with the cell-cycle regulation.
Intrathecal baclofen withdrawal syndrome- a life-threatening complication of baclofen pump: A case report
Authors: Mohammed Imran , Hussain Asif
Published: August 2004

Background

Intrathecal baclofen pump has been used effectively with increasing frequency in patients with severe spasticity, particularly for those patients who are unresponsive to conservative pharmacotherapy or develop intolerable side effects at therapeutic doses of oral baclofen. Drowsiness, nausea, headache, muscle weakness, light-headedness and return of pretreatment spasticity can be caused by intrathecal pump delivering an incorrect dose of baclofen. Intrathecal baclofen withdrawal syndrome is a very rare, potentially life-threatening complication of baclofen pump caused by an abrupt cessation of intrathecal baclofen.

Case presentation

A 24-year-old man with a past medical history of cerebral palsy and spastic quadriparesis developed hyperthermia, disseminated intravascular coagulation, rhabdomyolysis, acute renal failure and multisystem organ failure leading to a full-blown intrathecal baclofen withdrawal syndrome. Intrathecal baclofen pump analysis revealed that it was stopped due to some programming error. He was treated effectively with supportive care, high-dose benzodiazepines and reinstitution of baclofen pump.

Conclusion

The episodes of intrathecal baclofen withdrawal syndrome are mostly caused by preventable human errors or pump malfunction. Educating patients and their caregivers about the syndrome, and regular check-up of baclofen pump may decrease the incidence of intrathecal baclofen withdrawal syndrome. Oral baclofen replacement may not be an effective method to treat or prevent intrathecal baclofen withdrawal syndrome. Management includes an early recognition of syndrome, proper intensive care management, high-dose benzodiazepines and prompt analysis of intrathecal pump with reinstitution of baclofen.
Hereditary 1,25-Dihydroxyvitamin D Resistant Rickets due to a Mutation Causing Multiple Defects in Vitamin D Receptor Function
Authors: Malloy Peter J. , Xu Rong , Peng Lihong , Peleg Sara , Al-Ashwal Abdullah , Feldman David
Published: August 2004

Hereditary vitamin D-resistant rickets (HVDRR) is an autosomal recessive disease caused by mutations in the vitamin D receptor (VDR). We studied a young Saudi Arabian girl who exhibited the typical clinical features of HVDRR, but without alopecia. Analysis of her VDR gene revealed a homozygous T to C mutation in exon 7 that changed isoleucine to threonine at amino acid 268 (I268T). From crystallographic studies of the VDR ligand-binding domain, I268 directly interacts with 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] and is involved in the hydrophobic stabilization of helix H12. We recreated the I268T mutation and analyzed its effects on VDR function. In ligand binding assays, the I268T mutant VDR exhibited an approximately 5- to 10-fold lower affinity for [3H]1,25(OH)2D3 compared with the wild-type (WT) VDR. The I268T mutant required approximately a 65-fold higher concentration of 1,25(OH)2D3 to be equipotent in gene transactivation. Both retinoid X receptor heterodimerization and coactivator binding were reduced in the I268T mutant. Analogs of 1,25(OH)2D3 have been proposed as potential therapeutics for patients with HVDRR. Interestingly, in protease sensitivity assays, treatment with the potent vitamin D analog, 20-epi-1,25(OH)2D3, stabilized I268T mutant proteolytic fragments better than 1,25(OH)2D3. Moreover, 20-epi-1,25(OH)2D3 restored transactivation of the I268T mutant to levels exhibited by WT VDR treated with 1,25(OH)2D3. In conclusion, we describe a novel mutation, I268T, in the VDR ligand-binding domain that alters ligand binding, retinoid X receptor heterodimerization, and coactivator binding. These combined defects in VDR function cause resistance to 1,25(OH)2D3 action and result in the syndrome of HVDRR.
Role of Calcium-Calmodulin-Dependent Protein Kinase Cascade in Thyrotropin (TSH)-Releasing Hormone Induction of TSH and Prolactin Gene Expression
Authors: Murao Koji , Imachi Hitomi , Cao Wen M. , Yu Xiao , Tokumitsu Hiroshi , Inuzuka Hiroyuki , Wong Norman C. W. , Shupnik Margaret A. , Kobayashi Ryoji , Ishida Toshihiko
Published: August 2004

TRH binds to a membrane receptor that activates several intracellular signaling pathways and increases transcription of the TSH and prolactin (PRL) genes. Although TRH induces TSH and PRL gene expression, the underlying mechanism is not clear. In this report we examined the role of the Ca2+/calmodulin-dependent protein (CaM) kinase cascade in mediating TRH-stimulated transcription of TSH and PRL. RT-PCR and Western blot analysis were used to show that CaM kinase kinase (CaM-KK) and CaM IV (CaM-KIV) were present in rat anterior pituitary and its cell line GH3. Next, the effects of constitutively active CaM-KIV (CaM-KIVc) or its dominant negative mutant (CaM-KIVdn) on TSH and PRL promoter activity were tested in GH3 cells. The results showed that either CaM-KIVc alone or an upstream kinase, CaM-KK, induced the activity of both TSH and PRL promoters. Exposure of GH3 cells to 100 μm TRH induced CaM-KIV activity within 5 min and, as expected, also increased both TSH and PRL promoter activity. In contrast, cells carrying the CaM-KIVdn isoform had suppressed TRH induction of both TSH and PRL promoter activity. These results indicate that the CaM-KK-CaM-KIV cascade probably plays an important role in TRH induction of TSH and PRL transcriptional activity in pituitary cells.
Comparison of the Pharmacological Effects of a Novel Selective Androgen Receptor Modulator, the 5α-Reductase Inhibitor Finasteride, and the Antiandrogen Hydroxyflutamide in Intact Rats: New Approach for Benign Prostate Hyperplasia
Authors: Gao Wenqing , Kearbey Jeffrey D. , Nair Vipin A. , Chung Kiwon , Parlow A. F. , Miller Duane D. , Dalton James T.
Published: August 2004

Tissue-selective androgen receptor modulators (SARMs) demonstrate tissue selectivity in both castrated and intact male rats, behaving as partial agonists in androgenic tissues (i.e. prostate and seminal vesicle), but full agonists in anabolic tissues (i.e. levator ani muscle). The partial agonist activity of SARMs (compounds S-1 and S-4) in the prostate of intact rats suggested that SARM could be used for androgen suppression in the treatment of benign prostate hyperplasia (BPH). This study was designed to explore the mechanisms of action of SARM and to characterize the tissue selectivity of S-1 in intact male rats compared with that of hydroxyflutamide (antiandrogen) and finasteride (5α-reductase inhibitor), two major drugs used for androgen suppression treatment of BPH. In intact male rats, S-1 (5, 10, and 25 mg/kg) selectively decreased the prostate weight with similar efficacy to finasteride (5 mg/kg), without affecting the levator ani muscle or increasing the plasma levels of testosterone, LH, and FSH. Hydroxyflutamide (0.5, 1, 5, 10, and 25 mg/kg), however, decreased both the prostate and levator ani muscle weights without any selectivity and increased plasma hormone levels in a dose-dependent manner. Furthermore, S-1 and S-4 showed very weak inhibitory effects toward transiently expressed type I and II human 5α-reductase (Ki, >20 μm) during in vitro assays. Therefore, although S-1 and finasteride showed very similar suppressive effects in the prostate of intact male rats, they decreased prostate size via different mechanisms of action. S-1 simply worked as androgen receptor partial agonist, whereas finasteride inhibited prostatic 5α-reductase. These studies indicate that SARMs may demonstrate clinical utility as single agent or combination therapy for BPH.
Cytochrome P450-Dependent Lipid Metabolism in Preovulatory Follicles
Authors: Newman J. W. , Stok J. E. , Vidal J. D. , Corbin C. J. , Huang Q. , Hammock B. D. , Conley A. J.
Published: August 2004

Estrogen biosynthesis and proteolysis are both important processes involved in ovarian follicular development, which may be influenced by cytochrome P450 (CYP)-dependent fatty acid metabolites. However, CYP-dependent lipid metabolism has not been characterized with respect to follicular maturation in vivo. Therefore, follicular fluid was collected in the hours before and after the LH surge in pigs, and concentrations of epoxy, hydroxy, and dihydroxy lipids were measured by liquid chromatography tandem mass spectrometry. Arachidonate oxidation and epoxyeicosatrienoic acid hydrolysis to dihydroxyeicosatrienoic acids (DHETs) were also assessed in thecal and granulosa tissue fractions, and the expression of CYP epoxygenases was evaluated by immunoblots using available antisera. To evaluate soluble epoxide hydrolase (sEH) expression, the porcine sEH was cloned from ovarian tissue, expressed and purified for antibody generation. The follicular fluid oxylipin concentrations ranged from 1–150 nm depending on the compound and estrous stage. The follicular fluid concentrations of CYP-dependent oxylipins increased at estrus, as did sEH expression; however, significant changes in epoxides were not observed, and the 11,12-DHET peak was delayed. The ratio of 14,15–DHET:11,12-DHET across all samples correlated with the log of follicular fluid estradiol concentrations (P < 0.01). Epoxygenase activities were similar in theca and granulosa, varying little with follicular development, whereas the decline of a single CYP2J isoform at ovulation was observed by immunoblots. The sEH activity was higher in granulosa than in theca. Finally, the dynamic changes in follicular CYP-dependent arachidonic acid metabolites and their modulatory function in vascular models suggest roles for these metabolites in follicular maturation, which may include regulation of estradiol biosynthesis and preovulatory remodeling of the follicular wall that should be fully explored in future studies.
In the Adult Male Rhesus Monkey (Macaca mulatta), Unilateral Orchidectomy in the Face of Unchanging Gonadotropin Stimulation Results in Partial Compensation of Testosterone Secretion by the Remaining Testis
Authors: Simorangkir David R. , Ramaswamy Suresh , Marshall Gary R. , Plant Tony M.
Published: August 2004

This study examined, in adult monkeys, the role that gonadotropin-independent mechanisms play in compensation of testosterone (T) secretion by the testis that remains after unilateral orchidectomy (UO). We employed a model (testicular clamp), in which endogenous gonadotropin secretion was abolished with a GnRH receptor antagonist, and the gonadotropin drive to the testes was concomitantly replaced with an invariant iv pulsatile infusion of recombinant human LH and FSH (1-min pulse every 2.5 h: LH, 0.08–0.12 IU/kg·pulse; FSH, 0.12–0.32 IU/kg·pulse) that provided the Leydig cells with a physiological stimulus. Within 5 h of UO (n = 5), circulating T concentrations had declined to 43% of pre-UO levels. By d 4, however, loss of the first testis was partially compensated, as reflected by the finding that circulating T had reached a plateau of 67% of the pre-UO level, where it remained for the duration of the study (39 d). That the recovery in circulating T was the result of increased T secretion by the remaining testis was suggested by the finding that the pulsatile pattern and decay of T during the intergonadotropin pulse interval before and after UO were indistinguishable. Interestingly, inhibin B production by the remaining testis also showed a delayed, albeit, minor, compensation (13% on d 10–11; P > 0.05) after loss of the first testis. These results suggest that compensation in T production by the remaining testis after UO in adult monkeys may be achieved in part by a gonadotropin-independent mechanism that probably involves direct neural inputs to the primate testis.
Identity and Regulation of Stored and Secreted Progastrin-Derived Peptides in Sheep
Authors: Paterson Adrienne C. , Lockhart Sharon M. , Baker Josephine , Neumann Greg , Baldwin Graham S. , Shulkes Arthur
Published: August 2004

Amidated and nonamidated progastrin-derived peptides have distinct biological activities that are mediated by a range of receptor subtypes. The objective was to determine the nature of the stored and secreted progastrin-derived peptides and to investigate whether progastrin release is regulated by gastric acidity. Using an antiserum directed to the C terminus of progastrin for identification and to monitor purification, C-terminal flanking peptides (CTFP) of progastrin (prog76–83, prog77–83, and prog78–83 in approximately equivalent amounts) were isolated and identified from extracts of sheep antrum using ion exchange, HPLC, and mass spectrometry. Only trace amounts of full-length progastrin were present. Progastrin CTFP was the predominant progastrin-derived peptide in the antrum [progastrin CTFP/gastrin amide (Gamide) = 3]. Similarly, progastrin CTFP was the major circulating form in the antral (CTFP, 710 ± 62 pmol/liter; Gamide, 211 ± 35 pmol/liter) and jugular (CTFP, 308 ± 16 pmol/liter; gastrin amide, 32 ± 3 pmol/liter) veins. Alteration of gastric acidity in sheep by iv infusion of a H/K-adenosine triphosphatase inhibitor or somatostatin or by intragastric infusion of HCl demonstrated that the CTFP concentrations changed, although to a lesser extent than the changes in circulating gastrin amide. We conclude that the CTFP of progastrin is the major stored and circulating species of the gastrin gene, and that it is secreted in a regulated fashion rather than constitutively. Because full-length progastrin is bioactive, but is only a minor antral and secreted form, determination of the biological activity of the C-terminal flanking peptides will be important for a complete understanding of gastrin endocrinology.
Indirect Spectral Hard Modeling for the Analysis of Reactive and Interacting Mixtures
Authors: Alsmeyer Frank , Koß Hans-Jürgen , Marquardt Wolfgang
Published: August 2004

We present an indirect hard modeling (IHM) approach for the quantitative analysis of reactive multicomponent mixtures with intermolecular interaction. It can be used when it is not possible to obtain calibration data in the composition region of interest. The goal of this work, specifically, is to analyze reactive systems, although the validation of the method is done with nonreactive systems. Compared to conventional hard modeling, the new approach reduces the manual work required for modeling and renders unnecessary the assignment of bands in mixture spectra to individual components. It is based on parametric models of the pure component spectra that are made just flexible enough to fit the spectra of the unknown mixtures, and it only requires small calibration data sets that may lie in different regions of the composition space. The application to infrared (IR) and Raman spectra of multicomponent systems is discussed.
Resolution of Intermediate Adsorbate Structures in the Potential-Dependent Self-Assembly of n-Hexanethiolate on Silver by in situ Surface-Enhanced Raman Spectroscopy
Authors: Uibel Rory H. , Harris Joel M.
Published: August 2004

Resolution of the reaction steps and the associated component Raman spectra during the formation or desorption of self-assembled monolayers is challenging because intermediate adsorbate populations are present at low concentrations and their spectral bands overlap. By collecting Raman spectra versus applied potential into a two-dimensional data set, one can utilize multivariate statistical techniques to resolve the component concentration profiles along with their corresponding Raman spectra. In situ surface-enhanced Raman spectroscopy (SERS) spectra were collected during the potential-dependent formation and desorption (−1.50 to −0.70V versus Ag/AgCl) of n-hexanethiolate monolayer at a polycrystalline Ag electrode. Resolution of the pure component spectra from these components was accomplished by using self-modeling curve resolution (SMCR), which does not require a physical model. For monolayer adsorption, the potential-dependent Raman spectra could be described by three significant eigenvectors; the eigenvectors could be rotated into a set of pure component spectra and concentration profiles using a linear least-squares step to find a common plane in the space of the eigenvectors representing the linear combination of the real-component responses. The convex hull surrounding the data in the plane and positive amplitude criteria were utilized to identify the coordinates of the pure component responses. The C– S stretching vibrations of the resolved spectra show that the initial adsorbate is a gauche conformer, which allows the hydrocarbon chain to lie on the metal surface; a second phase arises at higher coverage with trans C–S conformation, where the hydrocarbon chains are oriented off the surface plane, and a final complete monolayer is formed with a well-ordered, all-trans C–S configuration. In contrast, desorption studies showed only two surface phases, the initial well-ordered monolayer and the low-density phase dominated by gauche conformations. The results illustrate the utility of self-modeling curve resolution to unravel interfacial reaction mechanisms and intermediate structures from two-dimensional SERS data, without requiring prior knowledge of a physical model for the process.
Gram—Schmidt Orthogonalization for Rapid Reconstructions of Fourier Transform Infrared Spectroscopic Imaging Data
Authors: Bhargava Rohit , Levin Ira W.
Published: August 2004

Increasingly voluminous Fourier transform infrared (FT-IR) spectroscopic imaging data sets are being generated with the advent of both faster array detectors and the implementation of time-resolved imaging techniques, resulting in data processing becoming the limiting step in visualizing sample heterogeneity and temporal profile evolution. We report the application of a Gram–Schmidt vector orthogonalization procedure in interferogram space to provide a significant time saving advantage in processing of one to two orders of magnitude in comparison to conventional spectral processing. Illustrative data from human skin biopsies and from dynamic molecular reorganizations within liquid crystalline microdomains is employed to discuss the capabilities and limitations of this information-extraction approach.
Automatic Generation of Peak-Shaped Models
Authors: Alsmeyer Frank , Marquardt Wolfgang
Published: August 2004

We describe how parametric spectral models for analytical applications can be generated by an automatic curve-fitting algorithm. The algorithm does not require initial choices of parameters or other human intervention, in contrast to established approaches that rely on deconvolution or derivative spectroscopy. This algorithm has been applied for quantitative analysis but can potentially be used in other applications that are based on parametric representations of peak-shaped models or could benefit from using such models, such as calibration transfer.
Chemical Segregation and Reduction of Raman Background Interference Using Drop Coating Deposition
Authors: Zhang Dongmao , Mrozek Melissa F. , Xie Yong , Ben-Amotz Dor
Published: August 2004

A new application of the recently described drop coating deposition Raman (DCDR) method facilitates the segregation and independent spectral characterization of mixture components. The quality of the normal (un-enhanced) Raman spectra are significantly improved as a result of reduced spectral interference from fluorescent impurities and buffer compounds. Fluorescence of commercial amino acid (O-phospho-L-serine) and protein (myoglobin) samples is reduced by over an order of magnitude using DCDR, more effectively than prolonged photo-bleaching. Furthermore, DCDR is used to obtain high-quality Raman spectra of proteins, lysozyme, and insulin, derived from solutions with up to 1000-fold excess buffer concentration. Possible thermodynamic and kinetic contributions to the observed segregation phenomena are discussed.
Simple Fiber-Optic-Based Sensors for Process Monitoring: An Application in Wine Quality Control Monitoring
Authors: Noiseux Isabelle , Long William , Cournoyer Alain , Vernon Marcia
Published: August 2004

The goal of this research was to develop a simple and economical fiber-optic sensor technology for agrifood process monitoring. Toward this end, two fiber-optic sensors were developed to be used in combination: a single reflection V-bend sensor and a single fiber air-gap probe. The former is designed to be sensitive toward refractive index and the latter towards absorption. Experiments indicate that the micromachined V-bend fiber refractometer is most sensitive when the bend angle is centered around 140 degrees, at which angle the sensor may resolve changes in refractive index as small as 0.00015. Additionally, the V-bend sensor was found to be non-responsive toward sample absorption even in extremely absorbing solutions. The air-gap design absorption sensor, most commonly used for measurements in highly colored media, was found to be slightly sensitive towards refractive index. When the two sensors are used together, the response of the absorption sensor may be corrected for. This sensor combination is able to provide accurate measurements in situations where Beer's law is not obeyed. Results are presented that show that the sensor pair was successfully used to monitor wine sugar content (Brix), and color density and hue, parameters related to the age of the wine.
Single-Tube Method for Nucleic Acid Extraction, Amplification, Purification, and Sequencing
Authors: Margraf Rebecca L , Page Sam , Erali Maria , Wittwer Carl T
Published: August 2004

Background: The hepatitis C virus (HCV) genotype determines patient prognosis and duration of treatment, but sequencing of the gene is lengthy and labor-intensive. We used a commercially available nucleic acid extraction system to develop a single-tube extraction-to-sequencing (STETS) method for HCV genotyping.

Methods: HCV RNA was purified and amplified in tubes coated with a solid-phase matrix that irreversibly bound nucleic acid during the extraction step. After reverse transcription-PCR, the amplicon was adsorbed to the original extraction matrix for purification and use in the subsequent sequencing reactions.

Results: The STETS method generated genotyping-quality sequence for a range of HCV titers from 500 to 6 000 000 IU/mL. If a viral sample was detected during real-time reverse transcription-PCR, it could be sequenced and genotyped. Read lengths >600 bases were observed with the STETS method. Mixed infections were detected and genotyped if at least 15% of the minor species was present. Combining the STETS method with consecutive sequencing provided a means of performing both forward and reverse sequencing in a single tube.

Conclusions: A single-tube nucleic acid extraction-to-sequencing method, which requires less time and labor than conventional methods, generates HCV sequence data that are equivalent to conventional methods and can be used to genotype HCV.
C-Reactive Protein and Features of the Metabolic Syndrome in a Population-Based Sample of Children and Adolescents
Authors: Lambert Marie , Delvin Edgard E , Paradis Gilles , O’Loughlin Jennifer , Hanley James A , Levy Emile
Published: August 2004

Background: C-Reactive protein (CRP) is a risk marker for type 2 diabetes and cardiovascular diseases. In youth, limited data are available on the distribution of high-sensitivity CRP as well as on its association with components of the metabolic syndrome.

Methods: In 1999, we conducted a school-based survey of a representative sample of youths 9, 13, and 16 years of age in the province of Quebec, Canada. Standardized clinical measurements and fasting plasma lipid, glucose, insulin, and CRP concentrations were available for 2224 individuals.

Results: The distribution of CRP was positively skewed. The median and 95th percentile values by age and sex ranged from <0.2 to 0.56 mg/L and from 2.72 to 6.28 mg/L, respectively. A total of 7.7% of 9-year-olds, 5.5% of 13-year-olds, and 12.8% of 16-year-olds had CRP concentrations >3.0 mg/L, the threshold defining the adult high-risk category. We observed a strong relationship between CRP concentrations and both body mass index (BMI) and fasting insulin values. The association between CRP and insulin concentration was markedly attenuated after adjustment for BMI, whereas that between CRP and BMI remained unchanged after adjustment for insulin: a 1 SD increase in BMI was associated with a 52% increase in CRP concentration. An increased CRP concentration was independently associated with a worsening of the lipid profile, whereas the association between increased CRP values and high systolic blood pressure was no longer statistically significant after adjustment for BMI.

Conclusions: The metabolic correlates of excess weight, including a state of low-grade systemic inflammation, are detectable early in life. Their health impact in adults remains to be fully examined.
Noninvasive Measurement of Glucose by Metabolic Heat Conformation Method
Authors: Cho Ok Kyung , Kim Yoon Ok , Mitsumaki Hiroshi , Kuwa Katsuhiko
Published: August 2004

Background: We developed a method, called the metabolic heat conformation (MHC) method, for the noninvasive measurement of blood glucose. The MHC method involves the measurement of physiologic indices related to metabolic heat generation and local oxygen supply, which correspond to the glucose concentration in the local blood supply.

Methods: We used noninvasive thermal and optical sensors on the fingertip of an individual to measure thermal generation, blood flow rate, hemoglobin (Hb) concentration, and oxyhemoglobin concentration. The calibration model incorporates mathematical procedures to convert signals from the sensor pickup to final glucose concentrations. The mathematical procedures are multivariate statistical analyses, involving values from sensor signals, polynomials from various values, regression analyses of individual patients, and cluster analyses of patient groups. The glucose value is calculated for each patient measurement, applying one of the clusters by discriminant analysis.

Results: Regression analysis was performed to compare the noninvasive method with the hexokinase method, using 127 data points (109 data points from diabetic patients, 18 data points from nondiabetic patients) with glucose concentrations ranging from 3.0 to 22.5 mmol/L (54–405 mg/dL). The correlation coefficient (r) was 0.91. Reproducibility was measured for healthy fasting persons; the CV was 6% at 5.56 mmol/L (100 mg/dL).

Conclusions: These data provide preliminary evidence that the MHC method can be used to estimate blood glucose concentrations noninvasively.
Diagnostic Probes for Bacillus anthracis Spores Selected from a Landscape Phage Library
Authors: Brigati Jennifer , Williams David D , Sorokulova Iryna B , Nanduri Viswaprakash , Chen I-Hsuan , Turnbough Charles L , Petrenko Valery A
Published: August 2004

Background: Recent use of Bacillus anthracis spores as a bioweapon has highlighted the need for a continuous monitoring system. Current monitoring systems rely on antibody-derived probes, which are not hardy enough to withstand long-term use under extreme conditions. We describe new, phage-derived probes that can be used as robust substitutes for antibodies.

Methods: From a landscape phage library with random octapeptides displayed on all copies of the major phage coat protein of the phage fd-tet, we selected clones that bound to spores of B. anthracis (Sterne strain). ELISA, micropanning, and coprecipitation assays were used to evaluate the specificity and selectivity with which these phage bound to B. anthracis spores.

Results: Peptides on the selected clones directed binding of the phage to B. anthracis spores. Most clones exhibited little or no binding to spores of distantly related Bacillus species, but some binding was observed with spores of closely related species. Our most specific spore-binding phage displayed a peptide EPRLSPHS (several thousand peptides per phage) and bound 3.5- to 70-fold better to spores of B. anthracis Sterne than to spores of other Bacillus species.

Conclusions: The selected phage probes bound preferentially to B. anthracis Sterne spores compared with other Bacillus species. These phage could possibly be further developed into highly specific and robust probes suitable for long-term use in continuous monitoring devices and biosorbents.
Functional genomics identifies novel and diverse molecular targets of nutrients in vivo
Authors: Gohil Kishorchandra
Published: August 2004

Genomic responses to nutrients are important determinants of physiological and pathological functions of living systems. Many of these responses are mediated by changes in mRNA concentrations that are primarily regulated by gene transcription. Transcriptional networks that regulate the expression and activities of transcription factors and structural genes in response to nutrients need to be defined. The tools of functional genomics and bioinformatics offer powerful means to address these needs. The application of global mRNA profiling tools to define genome-wide responses to nutrients and micronutrients with a primary focus on in vivo genomic responses of vital organs of laboratory mice is reviewed here. The studies show that major and minor nutrients affect the expression of mRNAs that are related to aging and inflammation, and chemically diverse micronutrients such as polyphenols and tocopherols may exert their effects through modulating the expression of functionally related genes.
Health Care for Children of Immigrant Families
Authors:   Committee on Community Health Services
Published: August 2004

The intent of this statement is to inform practitioners about the special health care needs and vulnerabilities of immigrant children and their families and to suggest clinical approaches to various aspects of their care. Immigrant children and their families, a large and diverse population group, have numerous risks to physical health and functioning and may be unfamiliar with our health care services. They often face many barriers to care, and their special risks and needs may not be familiar or readily apparent to many health care providers. Recently enacted federal welfare and immigration reform measures may increase the vulnerability of this population by limiting its access to health and social services. For multiple ethical and medical reasons, the American Academy of Pediatrics has historically opposed, and continues to oppose, denying needed services to any child residing within the borders of the United States.
Mycobacterium tuberculosis Transmission From a Pediatrician to Patients
Authors: Askew George L. , Finelli Lyn , Hutton Mary , Laraque Fabienne , Porterfield Debra , Shilkret Kenneth , Valway Sarah E. , Onorato Ida , Spitalny Kenneth
Published: August 2004

The following report describes the contact investigation of a pediatrician with tuberculosis (TB). The pediatrician's disease was discovered in late February 1993 after tuberculin skin testing (TST) of his 15-month-old son was positive (13-mm induration). Further investigation to identify the source of the child's infection revealed a positive (15-mm induration) TST in the pediatrician. The pediatrician had been symptomatic with a cough since September 1992. The pediatrician had a chest radiograph that revealed numerous cavitary lesions and a sputum smear that was positive for acid-fast bacilli. An investigation was initiated to assess whether the transmission of Mycobacterium tuberculosis had occurred in the pediatrician's office to patients, families, or other visitors. The investigation was later extended to include the hospitals and the day care center where the pediatrician worked.

Methods. A letter was mailed to parents of children served by the practice, explaining the potential exposure to TB and requesting that all persons who visited the office after September 1, 1992 complete an interview and Mantoux TST. Mass interviewing, testing, and test interpretation within the practice took place seven times during March and April 1993.

Results. At the completion of screening, 181 (87%) of 208 children who had close contact with the index case were reliably skin-tested and returned for interpretation. Three (1.7%) of the 181 children were TST-positive (≥5 mm). Thirty-seven (13%) of the 286 adults tested and returning for interpretations were TST-positive (≥10 mm). Thirty-two (86%) of the 37 adults who tested positive were foreign-born.

Conclusion. This investigation highlighted the need for identifying childhood TB infection as a sentinel event for adult disease. It also demonstrated the difficulty associated with deciding the extent of contact investigation of a health care worker with TB. Finally, the investigation emphasized the importance of maintaining regularly scheduled and appropriate testing for TB infection in health care workers and the need for health care workers to be cognizant of their own risk and be able to identify, especially in themselves, signs and symptoms of potential TB disease.
Comparison of Infasurf (Calf Lung Surfactant Extract) to Survanta (Beractant) in the Treatment and Prevention of Respiratory Distress Syndrome
Authors: Bloom Barry T. , Kattwinkel John , Hall Robert T. , Delmore Paula M. , Egan Edmund A. , Trout J. Richard , Malloy Michael H. , Brown David R. , Holzman Ian R. , Coghill Carl H. , Carlo Waldemar A. , Pramanik Arun K. , McCaffree Mary Anne , Toubas Paul L. , Laudert Susan , Gratny Linda L. , Weatherstone Kathleen B. , Seguin John H. , Willett Lynne D. , Gutcher Gary R. , Mueller Dawn H. , Topper William. H
Published: August 2004

Objective. To compare the relative safety and efficacy of Infasurf (calf lung surfactant extract; ONY, Inc, Amherst, NY, IND #27169) versus Survanta (Beractant, Ross Laboratories, Columbus, OH) in reducing the acute severity of respiratory distress syndrome (RDS) when given at birth and to infants with established RDS.

Design. A prospective, randomized, double-blind, multicenter clinical trial.

Setting. Thirteen neonatal intensive care units participated in the treatment arm: seven of these concurrently participated in the prevention arm.

Patients. The treatment arm enrolled infants of ≤2000 g birth weight with established RDS, and the prevention arm enrolled infants of ≤29 weeks' gestation with birth weights <1250 g.

Intervention. Infants were randomly assigned to receive Infasurf (n = 303, treatment arm; n = 180, prevention arm) or Survanta (n = 305, treatment arm; n = 194, prevention arm) in accordance with the Survanta package insert instructions.

Outcome Measures. We projected a 25% reduction between groups in the need for a third dose of surfactant for infants with established RDS, and a 25% reduction in the need for a second dose of surfactant for infants who received prophylactic surfactant. Secondary outcomes included the severity of RDS measured by inspired oxygen concentrations and mean airway pressure, air leaks, complications associated with surfactant administration, and survival to 36 weeks' postmenstrual age without the need for oxygen supplementation.

Results. In the treatment arm, there was no difference between groups in the number of infants requiring more than two doses of surfactant. The interval between doses was significantly longer for Infasurf, suggesting an increased duration of treatment effect. The inspired oxygen concentration and mean airway pressure were lower in the Infasurf infants during the first 48 hours in the treatment arm.

In the prevention arm, there were no differences with respect to the number of surfactant doses. The dosing intervals were longer for Infasurf infants after the second dose. No difference in inspired oxygen or mean airway pressure was noted during the first 72 hours.

There were no significant differences in the incidence of air leaks, complications associated with dosing, complications of prematurity, mortality, or survival without chronic lung disease in the prevention or treatment arm.

Conclusions. Infants treated with Infasurf have a modest benefit in the acute phase of RDS. Infasurf seems to produce a longer duration of effect than Survanta.
Early Discharge From the Newborn Nursery—Effect on Scheduling of Follow-up Visits by Pediatricians
Authors: Maisels M. Jeffrey , Kring Elizabeth
Published: August 2004

Objective. To evaluate the impact of shorter hospital stays on the follow-up scheduling of newborn infants by private pediatricians.

Design. Five surveys over a period of 18 months with educational intervention.

Setting. Large community hospital well baby nursery.

Participants. Twenty private pediatricians who cared for at least 20 newborn infants in the well baby nurseries during 1995.

Intervention. Oral and written communications to pediatricians emphasizing the importance of evaluating infants within 2 to 3 days of discharge if the hospital stay was less than 48 hours.

Main outcome measure. Interval between discharge from the nursery and the scheduled follow-up visit to the pediatrician.

Results. In the first two surveys (September 1994 and March 1995) there was no significant difference in follow-up scheduling by pediatricians for those infants discharged <48 hours vs ≥48 hours. Differences were significant in July and November 1995, and in the final survey in March 1996. Nevertheless, in March 1996, 38% of short-stay infants were scheduled to be seen 4 or more days after discharge, and 33% 14 days after discharge.

Conclusion. Although follow-up practices have changed in response to shorter newborn hospital stays, a significant proportion of pediatricians are not following the American Academy of Pediatrics guidelines for the follow-up of short-stay infants. Whether or not failure to follow these guidelines will lead to an increase in morbidity is unknown.
Human Monocytic Ehrlichiosis in Children
Authors: Schutze Gordon E. , Jacobs Richard F.
Published: August 2004

Background. Much of what is known about human monocytic ehrlichiosis (HME) is based upon studies with adult patients.

Purpose. To review our experience with HME to better understand the epidemiology, clinical manifestations, and outcome of this disease in children.

Methods. Demographic, clinical, and laboratory data were gathered after review of the medical records of patients identified with HME.

Results. Twelve patients with an median age of 7.4 years (range, 7 months to 13.7 years) were identified with HME; 10 were white, 7 were male, and 10 were from hometowns of <800 people. Eight patients presented from May through July, and 8 had a history of tick bites. Symptoms demonstrated by the patients during their illness included fever (100%), rash (67%), myalgias (58%), and vomiting, diarrhea, and headache (25%). On presentation, patients demonstrated thrombocytopenia (92%), elevated liver function tests (91%), lymphopenia (75%), hyponatremia (67%), leukopenia (58%), and anemia (42%) on the initial laboratory examination. Four patients presented in shock and 3 required blood pressure support and mechanical ventilation for a median of 10 days (8 to 37 days). These complicated patients required longer hospitalization (19.5 days vs 5.5 days) and attained higher blood urea nitrogen levels (42.5 mg/dL vs 10 mg/dL) than the patients not presenting with shock. Morbidity associated with HME patients included a decrease in cognitive and neurologic performance.

Conclusions. More information and long-term follow-up is required to understand the full spectrum of disease and morbidity associated with HME in children.
Surgical Treatment of Craniosynostosis: Outcome Analysis of 250 Consecutive Patients
Authors: Sloan Gerald M. , Wells Karin C. , Raffel Corey , McComb J. Gordon
Published: August 2004

Objective. Surgery for craniosynostosis has evolved rapidly over the past two decades, with increased emphasis on early, extensive operations. Older published series may not accurately reflect more recent experience. Our study was designed to analyze outcome in a large series of consecutive patients treated recently at a single center.

Methods. We reviewed 250 consecutive patients who underwent surgical treatment of craniosynostosis between January 1, 1987 and December 31, 1992. They were divided into nine groups by suture involvement: sagittal, unilateral coronal, bilateral coronal, unilateral lambdoid, bilateral lambdoid, metopic, multiple suture, the Klee-blattschädel deformity (cloverleaf skull), and acquired craniosynostosis. Outcome was analyzed in terms of residual deformities and irregularities, complications, mortality, as well as the need for additional surgery.

Results. There were 157 males (62.8%) and 93 females (37.2%), with most of the male preponderance accounted for by the large sagittal synostosis group, which consisted of 82 males and 25 females. Median age at first operation was 147 days. A named syndrome was present in 23 patients (9.2%) and was more common than expected with bilateral and unilateral coronal synostosis, the Kleeblattschädel deformity, and multiple suture synostosis. There were two deaths (0.8%), both with Klee-blattschädel patients, and 17 other complications (6.8%). Morbidity and mortality were significantly associated with secondary vs primary operations and syndromic vs nonsyndromic patients. Outcome analysis revealed the best surgical results with metopic synostosis and significantly less good results with the Kleeblattschädel deformity, multiple suture synostosis, and bilateral coronal synostosis.

Conclusions. Using modern surgical techniques, craniosynostosis can be corrected with good outcomes and relatively low morbidity and mortality, particularly for otherwise healthy, nonsyndromic infants.