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Biomedical subjects

J Klein

Publications and source records attributed to J Klein.

At least 109 records · Page 6Linked to original sources
PubMed2001

Towards functional proteomics of membrane protein complexes: analysis of thylakoid membranes from Chlamydomonas reinhardtii.

Functional proteomics of membrane proteins is an important tool for the understanding of protein networks in biological membranes but structural studies on this part of the proteome are limited. In this study we undertook such an approach to analyse photosynthetic thylakoid membranes isolated from wild-type and mutant strains of Chlamydomonas reinhardtii. Thylakoid membrane proteins were separated by high-resolution two-dimensional gel electrophoresis (2-DE) and analysed by immuno-blotting and mass spectrometry for the presence of membrane-spanning proteins. Our data show that light-harvesting complex proteins (LHCP), that cross the membrane with three transmembrane domains, can be separated using this method. We have identified more than 30 different LHCP spots on our gels. Mass spectrometric analysis of 2-DE separated Lhcb1 indicates that this major LHCII protein can associate with the thylakoid membrane with part of its putative transit sequence. Separation of isolated photosystem I (PSI) complexes by 2-DE revealed the presence of 18 LHCI protein spots. The use of two peptide-specific antibodies directed against LHCI subunits supports the interpretation that some of these spots represent products arising from differential processing and post-translational modifications. In addition our data indicate that the reaction centre subunit of PSI, PsaA, that possesses 11 transmembrane domains, can be separated by 2-DE. Comparison between 2-DE maps from thylakoid membrane proteins isolated from a PSI-deficient (Deltaycf4) and a crd1 mutant, which is conditionally reduced in PSI and LHCI under copper-deficiency, showed the presence of most of the LHCI spots in the former but their absence in the latter. Our data demonstrate that (i) hydrophobic membrane proteins like the LHCPs can be faithfully separated by 2-DE, and (ii) that high-resolution 2-DE facilitates the comparative analysis of membrane protein complexes in wild-type and mutants cells.

Amino Acid Sequence↗
PubMed2001

Compensatory mechanisms enhance hippocampal acetylcholine release in transgenic mice expressing human acetylcholinesterase.

Central cholinergic neurotransmission was studied in learning-impaired transgenic mice expressing human acetylcholinesterase (hAChE-Tg). Total catalytic activity of AChE was approximately twofold higher in synaptosomes from hippocampus, striatum and cortex of hAChE-Tg mice as compared with controls (FVB/N mice). Extracellular acetylcholine (ACh) levels in the hippocampus, monitored by microdialysis in the absence or presence of 10(-8)-10(-3) M neostigmine in the perfusion fluid, were indistinguishable in freely moving control and hAChE-Tg mice. Muscarinic receptor functions were unchanged as indicated by similar effects of scopolamine on ACh release and of carbachol on inositol phosphate formation. However, when the mice were anaesthetized with halothane (0.8 vol. %), hippocampal ACh reached significantly lower levels in AChE-Tg mice as compared with controls. Also, the high-affinity choline uptake (HACU) in hippocampal synaptosomes from awake hAChE-Tg mice was accelerated but was reduced by halothane anaesthesia. Moreover, hAChE-Tg mice displayed increased motor activity in novel but not in familiar environment and presented reduced anxiety in the elevated plus-maze test. Systemic application of a low dose of physostigmine (100 microgram/kg i.p.) normalized all of the enhanced parameters in hAChE-Tg mice: spontaneous motor activity, hippocampal ACh efflux and hippocampal HACU, attributing these parameters to the hypocholinergic state due to excessive AChE activity. We conclude that, in hAChE-Tg mice, hippocampal ACh release is up-regulated in response to external stimuli thereby facilitating cholinergic neurotransmission. Such compensatory phenomena most likely play important roles in counteracting functional deficits in mammals with central cholinergic dysfunctions.

Acetylcholine↗
PubMed2001

Fatigue and hyponatremia in a 75-year-old woman: unusual presentation of hypophysitis.

A 75-year-old woman presented with general fatigue progressing to somnolence. Laboratory tests showed marked hyponatremia. TSH in the normal range, but low levels of free T3 and free T4. Evaluation of pituitary hormones and magnetic resonance imaging of the pituitary unmasked findings characteristic for hypophysitis with secondary adrenal insufficiency and secondary hypothyroidism. Hormonal substitution with hydrocortisone and levothyroxine resulted in rapid improvement of all symptoms and signs. Without additional treatment shrinkage of the pituitary gland could be documented. Our report extends the known clinical and pathological spectrum of hypophysitis and illustrates the need to include this uncommon entity in the differential diagnosis of hyponatremia even in elderly patients.

Adrenal Insufficiency↗
PubMed2001

Parapneumonic effusions and empyema.

Parapneumonic effusions occur in up to 60% of patients hospitalized with community acquired pneumonia. Fortunately, the majority of these effusions follow an uncomplicated course responding to antibiotic therapy alone. An important subgroup of patients, however, follow a complicated course progressing to an empyema unless infected pleural fluid is drained. Decisions for drainage depend on host factors, the nature of the respiratory pathogen, and the extent of pleural inflammation and loculation. Staging of a parapneumonic effusion allows selection of a drainage procedure that will be most effective. Adjunctive therapies, such as intrapleural fibrinolysis, appear to contribute to resolution of pleural infection but more investigational data are needed to determine their specific role.

Journal Article↗
PubMed2001

Assessing fertility in women of advanced reproductive age.

Reproductive capacity in women declines dramatically beyond the fourth decade of life. Oocyte quality seems to be the primary determinant of reproductive potential, although age-related uterine changes may also contribute. Underlying reasons for reproductive decline in women remain unclear, and both ovarian and neuroendocrine mechanisms have been proposed.A number of age-related endocrinologic changes precede menopause and predict diminished reproductive capacity. Thus, "ovarian reserve" screening may identify patients in whom attempts at conventional assisted reproduction is warranted before proceeding with ovum donation. Techniques to preserve the maternal genetic contribution to offspring (in lieu of egg donation), including germinal vesicle transfer and donor ooplasm, are under investigation.

Adult↗
PubMed2001

Use of hair analysis for confirmation of self-reported cocaine use in users with negative urine tests.

INTRODUCTION: Identification of cocaine use based on a urine test may miss many cases because of the short elimination half-life of the drug. Our objective was to verify the sensitivity of the cocaine hair test in admitted users. PATIENTS AND METHODS: Admitted cocaine users (38), that were 18-70 years of age and reported to have refrained from using cocaine in the few days to months prior to the test, were compared to 10 controls who claimed never to have used cocaine. All had negative urine tests for cocaine and benzoylecgonine by thin-layer chromatography. Cocaine and benzoylecgonine were extracted from unwashed hair and tested by established immunoassays. RESULTS: The hair test was positive in 37/38 cases (97%) and in none of the controls. There was significantly more cocaine in black hair than in brown or blonde hair per mg of cocaine dose reported to have been consumed, highlighting a potential bias when interpreting test results in individuals with dark hair. There was a statistically significant correlation between reported dose used and hair concentrations of cocaine. DISCUSSION: The cocaine hair test appears to be highly sensitive and specific in identifying past cocaine use in the setting of a negative urine test.

Adolescent↗
PubMed2001

On the origin of Darwin's finches.

Darwin's finches comprise a group of 15 species endemic to the Galápagos (14 species) and Cocos (1 species) Islands in the Pacific Ocean. The group is monophyletic and originated from an ancestral species that reached the Galápagos Archipelago from Central or South America. Descendants of this ancestor on the Archipelago then colonized Cocos Island. In the present study, we used sequences of two mitochondrial (mt) DNA segments (922 bp of the cytochrome b gene and 1,082 bp of the control region), as well as two nuclear markers (830 bp of numt2, consisting of 140 bp of mtDNA control region and 690 bp of flanking nuclear DNA; and 740 bp of numt3, consisting of 420 bp of mt cytochrome b sequence flanked by 320 bp of nuclear DNA) to identify the species group most closely related to the Darwin's finches. To this end, we analyzed the sequences of 28 species representing the main groups (tribes) of the family Fringillidae, as well as 2 outgroup species and 13 species of Darwin's finches. In addition, we used mtDNA cytochrome b sequences of some 180 additional Fringillidae species from the database for phylogeny reconstruction by maximum-parsimony, maximum-likelihood, minimum-evolution, and neighbor-joining methods. The study identifies the grassquit genus Tiaris, and specifically the species Tiaris obscura, as the nearest living relative of Darwin's finches among the species surveyed. Darwin's finches diverged from the Tiaris group shortly after the various extant species of Tiaris diverged from one another. The initial adaptive radiation of the Tiaris group apparently occurred on the Caribbean islands and then spread to Central and South America, from where the ancestors of Darwin's finches departed for the Galápagos Islands approximately 2.3 MYA, at the time of the dramatic climatic changes associated with the closure of the Panamanian isthmus and the onset of Pleistocene glaciation.

Animals↗
PubMed2001

Auditory information processing during adequate propofol anesthesia monitored by electroencephalogram bispectral index.

UNLABELLED: Memory for intraoperative events may arise from inadequate anesthesia when the hypnotic state is not continuously monitored. Electroencephalogram bispectral index (BIS) enables monitoring of the hypnotic state and titration of anesthesia to an adequate level (BIS 40 to 60). At this level, preserved memory function has been observed in trauma patients. We investigated memory formation in elective surgical outpatients during target-controlled propofol anesthesia supplemented with alfentanil. While BIS remained between 40 and 60, patients listened to a tape with either familiar instances (exemplars) from two categories (Experimental [E] group, n = 41) or bird sounds (Control [C] group, n = 41). After recovery, memory was tested directly and indirectly. BIS during audio presentation was on average (+/- SD) 44 +/- 5 and 46 +/- 5 for Groups E and C, respectively. No patient consciously recalled the intraoperative period, nor were presented words recognized reliably (Group E, 0.9 +/- 0.8 hits; Group C, 0.8 +/- 0.8 hits) (P = 0.7). When asked to generate category exemplars, Group E named 2.10 +/- 1.0 hits versus 1.98 +/- 1.0 in Group C (P = 0.9). We found no explicit or implicit memory effect of familiar words presented during adequate propofol anesthesia at BIS levels between 40 and 60 in elective surgical patients. IMPLICATIONS: This study suggests that stable levels of adequate hypnosis may prevent information processing and memory formation during general anesthesia and supports the feasibility of electroencephalogram bispectral index as a monitor of adequate anesthesia.

Acoustic Stimulation↗
PubMed2001

Assessing health system provision of adolescent preventive services: the Young Adult Health Care Survey.

BACKGROUND: Adolescents often do not receive recommended preventive counseling and screening services. Few measures are available to assess health care system performance in this area. OBJECTIVE: Develop a reliable, valid, and feasible method for measuring adherence to consensus guidelines for adolescent preventive counseling and screening services. METHODS: The 45-item Young Adult Health Care Survey (YAHCS) was tested with a diverse group of commercially and publicly insured adolescents enrolled in managed care organizations (n = 4,060). Psychometric, bivariate, and multivariate analyses were conducted to assess the reliability, validity, and patterns of variation in the preventive care measurement scales derived from the YAHCS. RESULTS: YAHCS measurement scales demonstrated strong construct validity (mean factor loading = 0.64) and reliability (mean Cronbach's alpha = 0.77). Average preventive counseling and screening scores ranged from 18.2% for discussing risky behavior topics to 50.4% for discussing diet, weight, and exercise topics. Adolescent demographic, health care use, and payer factors explained a small amount of variation across adolescent scores on YACHS scales (mean R2 = 0.086). Females and older teens were more likely to report private time with providers and counseling and screening on topics related to sex. Overall, the odds of receiving preventive counseling and screening for adolescents who reported having private time with providers, engaging in risky behaviors, or both were higher than for adolescents who did not meet privately or report risky behaviors (private visit OR, 3.60; 95% CI, 2.91-4.47; risky behaviors OR, 2.02; 95% CI, 1.62-2.52). CONCLUSIONS: The YAHCS provides a feasible, reliable, and valid method for assessing adherence to adolescent preventive services guidelines. It differentiates among varied aspects of preventive care provided to adolescents and is promising as a potential measure of health plan and provider quality. Improved performance on the YAHCS would indicate progress toward the achievement of Healthy People 2010 goals.

Adolescent↗
PubMed2001

Heavily smoking women who cannot quit in pregnancy: evidence of pharmacokinetic predisposition.

SUMMARY: Smoking in pregnancy is associated with a well-characterized increase in perinatal risks. Despite their wish to discontinue smoking, some pregnant women cannot stop. To characterize nicotine and cotinine levels in women who could not quit smoking after the first trimester, the authors recruited 19 white women (age 17-41 years) between 14-23 weeks of gestation who could not quit smoking. They started smoking at ages 11-22 years (mean 14.5) and smoked for 17 +/- 6 years. They had their first cigarettes 5-60 minutes after waking up (mean 12). Nicotine levels were compared with those expected in white patients in the general population, and the cotinine levels per cigarette smoked were compared with the population-based values. Sixteen of the 19 women had nicotine levels substantially lower than those expected. The mean level of serum cotinine produced by one cigarette per day was 19.1 +/- 15.8 ng/mL (range 6.1-67). The expected levels in white patients in the general population are 13 +/- 7.7 ng/mL. The data suggest that pregnant women who cannot quit heavy smoking in the second trimester form a selective group with pharmacokinetic predisposition to a high rate of nicotine metabolism.

Adolescent↗
PubMed2001

Glutathione stability in whole blood: effects of various deproteinizing acids.

High-performance liquid chromatography separation of reduced and oxidized glutathione (GSH and GSSG) in biologic samples using electrochemical detection offers the convenience of both simultaneous quantitation and simple sample preparation. Rapid acidification is required to prevent GSH autooxidation, GSH and GSSG degradation, and precipitate proteins that interfere with analysis. Currently, little consistency exists in the literature regarding acid selection or the feasibility of sample storage before analysis. The purpose of this work was to examine the effects of perchloric (PCA), trichloroacetic (TCA), metaphosphoric (MPA), and 5-sulfosalicylic (SSA) acids on the short-term stability of GSH and GSSG measurements in whole blood. Samples were collected from adult volunteers and treated with multiple concentrations of each acid. The samples were analyzed immediately and aliquots were stored at -80 degrees C for up to 28 days. The suitability of each acid was assessed by percentage change of GSH and GSSG from baseline, efficiency of protein removal, and alteration of chromatogram characteristics. In general, increasing the acid concentration improved sample stability. Nevertheless, SSA did not achieve acceptable sample stability at any concentration tested. MPA was found to leave substantial amounts of protein in the samples, and TCA may interfere with the peaks of interest. Based on these results, a final concentration of 15% PCA is suggested for analysis of glutathione in whole blood. Although immediate sample preparation is preferred, 15% PCA can maintain sample integrity for 4 weeks after storage at -80 degrees C.

Adult↗
PubMed2001

Pro-inflammatory cytokine production by synoviocytes following exposure to protein I/II, a modulin from oral streptococci.

We have tested the ability of protein I/II, an adhesin from oral streptococci, to stimulate the production of pro-inflammatory cytokines by synovial cells isolated from both rheumatoid arthritis and control patients. Protein I/II triggers synovial fluid cells to produce interleukin (IL)-6 and IL-8 while secretion of tumor necrosis factor-alpha (TNF-alpha) was less enhanced. Using fibroblast-like synoviocytes, we found that protein I/II also exerts an immunomodulatory effect (IL-6 and IL-8 release) on these cells. These findings indicate that, if it gains access to the joint cavity, protein I/II could participate in the initiation and/or perpetuation of rheumatic diseases, by stimulating pro-inflammatory cytokine release from various synovial cells.

Adhesins, Bacterial↗
PubMed2001

Essential role of insulin receptor substrate 1 in differentiation of brown adipocytes.

The most widely distributed members of the family of insulin receptor substrate (IRS) proteins are IRS-1 and IRS-2. These proteins participate in insulin and insulin-like growth factor 1 signaling, as well as the actions of some cytokines, growth hormone, and prolactin. To more precisely define the specific role of IRS-1 in adipocyte biology, we established brown adipocyte cell lines from wild-type and IRS-1 knockout (KO) animals. Using differentiation protocols, both with and without insulin, preadipocyte cell lines derived from IRS-1 KO mice exhibited a marked decrease in differentiation and lipid accumulation (10 to 40%) compared to wild-type cells (90 to 100%). Furthermore, IRS-1 KO cells showed decreased expression of adipogenic marker proteins, such as peroxisome proliferator-activated receptor gamma (PPARgamma), CCAAT/enhancer-binding protein alpha (C/EBPalpha), fatty acid synthase, uncoupling protein-1, and glucose transporter 4. The differentiation deficit in the KO cells could be reversed almost completely by retrovirus-mediated reexpression of IRS-1, PPARgamma, or C/EBPalpha but not the thiazolidinedione troglitazone. Phosphatidylinositol 3-kinase (PI 3-kinase) assays performed at various stages of the differentiation process revealed a strong and transient activation in IRS-1, IRS-2, and phosphotyrosine-associated PI 3-kinase in the wild-type cells, whereas the IRS-1 KO cells showed impaired phosphotyrosine-associated PI 3-kinase activation, all of which was associated with IRS-2. Akt phosphorylation was reduced in parallel with the total PI 3-kinase activity. Inhibition of PI 3-kinase with LY294002 blocked differentiation of wild-type cells. Thus, IRS-1 appears to be an important mediator of brown adipocyte maturation. Furthermore, this signaling molecule appears to exert its unique role in the differentiation process via activation of PI 3-kinase and its downstream target, Akt, and is upstream of the effects of PPARgamma and C/EBPalpha.

Adipocytes↗
PubMed2001

Management of spontaneous pneumothorax: an American College of Chest Physicians Delphi consensus statement.

OBJECTIVE: Provide explicit expert-based consensus recommendations for the management of adults with primary and secondary spontaneous pneumothoraces in an emergency department and inpatient hospital setting. The use of opinion was made explicit by employing a structured questionnaire, appropriateness scores, and consensus scores with a Delphi technique. The guideline was designed to be relevant to physicians who make management decisions for the care of patients with pneumothorax. OPTIONS: Decisions for observation, chest tube placement, surgical interventions, and radiographic imaging. OUTCOMES: Effectiveness of pneumothorax resolution, duration of and patient tolerance of care, and pneumothorax recurrence. EVIDENCE: Literature review from 1967 to January 1999 and Delphi questionnaire submitted in three iterations to a multidisciplinary physician panel. VALUES: The guideline development group determined by consensus the relevant outcomes to be considered in developing the Delphi questionnaire. BENEFITS, HARMS, AND COSTS: The type and magnitude of benefits, harms, and costs expected for patients from guideline implementation. RECOMMENDATIONS: Management decisions vary between patients with primary or secondary pneumothoraces, with observation of small pneumothoraces being appropriate only for primary pneumothoraces. The level of consensus varies regarding the specific interventions indicated, but agreement exists for the general principles of care. VALIDATION: Recommendations were peer reviewed by physician experts and were reviewed by the American College of Chest Physicians (ACCP) Health and Science Policy Committee. IMPLEMENTATION: The guideline recommendations will be published in printed and electronic form with distribution of synopses for patients and health care providers. Contents of the guideline will be incorporated into continuing medical education programs. SPONSORS: The ACCP.

Adult↗
PubMed2001

High-level bacterial expression of a natively folded, soluble extracellular domain fusion protein of the human luteinizing hormone/chorionic gonadotropin receptor in the cytoplasm of Escherichia coli.

We have expressed the extracellular domain of the human luteinizing hormone/chorionic gonadotropin (hLH/CG) receptor as a fusion protein with thioredoxin in the cytoplasm of an Escherichia coli strain that contains mutations in both the thioredoxin reductase and glutathione reductase genes. The chimeric protein isolated following induction of expression is purified in a soluble form and binds hCG with an affinity approximating that of native receptor. This truncated form of the receptor displays the same specificity as intact hLH/CG receptor and does not bind human follicle stimulating hormone. This cytoplasmically produced protein is expressed at levels that exceed 10 mg/L. Expression of properly folded extracellular domain of the hLH/CG receptor in the cytoplasm of E. coli allows the facile and economic purification of large quantities of material. This will facilitate the determination of the structure of the hormone-binding domain of the glycoprotein receptor as well as the production of epitope-specific antibodies.

Amino Acid Sequence↗
PubMed2001

[Effect of portosystemic anastomosis on the degree of splenic inhibition].

UNLABELLED: Secondary hypersplenism is a well-known complication of portal hypertension. Reduced platelet count is a more alarming sign for the physician than risk for the patient. Improvement of thrombocytopenia is urgent, when portal hypertension with splenomegaly and thrombocytopenia presents with life-threatening haemorrhage from gastroesophageal varices. In this case, treatment aimed at stopping the bleeding may be more beneficial than any intervention on the spleen. In this study, we evaluated long-term effects of an elective distal splenorenal shunt or small diameter H-shunt on splenomegaly and thrombocytopenia in 26 patients with portal hypertension operated for repeated bleeding from oesophageal varices. 25 patients had splenomegaly and 16 patients had thrombocytopenia before shunting. Surgery corrected splenomegaly in 16 patients (64%), platelet counts increased in 13 of 16 patients with thrombocytopenia (81.2%). CONCLUSION: Selective or partial portal decompression is sufficient to alleviate thrombocytopenia and splenomegaly associated with portal hypertension.

Adult↗