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H Cramer

Publications and source records attributed to H Cramer.

At least 37 records · Page 2Linked to original sources
PubMed1998

Preparation and chromatographic purification of 125I-[Tyr0]-somatostatin-14 for the use in radioimmunoassay and receptor-binding experiments.

[Tyr0]-somatostatin-14 (SST-14) was converted to the corresponding radiolabeled 125I-[Tyr0]-SST-14 derivative by use of the chloramine-T technique. After solid-phase extraction (SPE) of the crude radiopeptide with microfine silica gel and desorption with acetone-water-acetic acid 39:40:1, the label was subjected to size exclusion chromatography (SEC) on Sephadex G-25 fine. Fractions attributable to the target compound were collected and investigated with respect to their specific as well as non-specific binding to a specific anti-somatostatin antibody. All fractions exhibiting an optimum ratio of specific versus non-specific binding were pooled and lyophilized for their further use in both radioimmunoassay (RIA) measurements of somatostatin like immunoreactivity (SLI) and receptor-binding experiments. A specific activity of approx. 1.6 x 10(6) Ci/M was calculated for the radiolabel prepared in this manner. Approximately 85-90% of radioactivity attributable to labeled [Tyr0]-SST species was incorporated into the desired mono-iodinated component. When 125I-[Tyr0]-SST-14 was purified by reversed-phase high performance liquid chromatography (RP-HPLC) using isocratic elution with 0.1 M triethylammonium formate (TEAF) buffer of pH 2.2 in 22% acetonitrile after prior SPE, specific binding decreases to about 80% compared with the value obtained for the radiopeptide subjected to SEC. Nevertheless, RP-HPLC proves as an efficient tool for rapid purity control of 125I-[Tyr0]-somatostatin-14 samples at different storage time intervals.

Animals↗
PubMed1998

Subtype-specific endothelin-A and endothelin-B receptor desensitization correlates with differential receptor phosphorylation.

In the rat cardiovascular system endothelin-1 (ET-1) elicits prolonged physiologic responses mediated by the ETA receptor, whereas the effects mediated by the ETB receptor are transient. The molecular mechanisms for the subtype-specific responses are not yet clear. However, post-translational modifications such as phosphorylation and palmitoylation may play an important role. In Sf9 cells overexpressing the human ETA and ETB receptors, both subtypes are palmitoylated. However, only the ETB but not the ETA receptor is phosphorylated in a ligand-dependent manner. Because phosphorylation is believed to play an important role in ligand-dependent receptor inactivation, we analyzed whether the differential phosphorylation of the ETA and ETB receptors reflects a differential mechanism of receptor inactivation. Using a modified inositol phosphate accumulation assay, we analyzed CHO cells that expressed the ETA or ETB receptor. The ETB receptor was deactivated almost completely within 5 min after agonist stimulation, whereas stimulation of the ETA receptor resulted in sustained activation, i.e., > 90% of the initial activity was maintained after 5 min of ligand stimulation and > 30% after 20 min. A strong correlation was observed between the time course of ETA receptor inactivation and ETA receptor internalization. The endogenous ETA receptor in Rat-1 cells produced a prolonged stimulation of phospholipase C similar to that seen in CHO cells. Therefore, the sustained signaling activity of the ETA receptor is not a property only of recombinant cell lines. Together, our data suggest rapid ETB receptor inactivation due to phosphorylation and delayed ETA receptor inactivation by internalization. These mechanisms adequately reflect the differential response patterns of the ET receptors under physiologic conditions.

Animals↗
PubMed1997

The AutoPap 300 QC System multicenter clinical trials for use in quality control rescreening of cervical smears: I. A prospective intended use study.

BACKGROUND: The AutoPap 300 QC System is an automated device for the analysis of conventionally prepared cervical cytology slides. The AutoPap System selects an enriched population of cases for human quality control (QC) review. The device assigns a score based on the likelihood that a slide is abnormal. Cases are selected for QC rescreening that have scores exceeding a preset threshold corresponding to approximately the top 10% (or greater) of scores. METHODS: AutoPap false-negative detection, compared with a 10% random QC process, was tested in a 6-center clinical evaluation study. At each site, a block of up to 150 consecutive negative slides (including detected false-negative cases) were selected randomly daily. All slides were run on the AutoPap System and rescreened by cytotechnologists for truth determination. The false-negative cases included in the top 10% group selected by AutoPap System then were compared with false-negative detection by the random selection process. RESULTS: Fourteen thousand nine hundred fourteen cases were analyzed. The AutoPap-enriched 10% quality control group contained false-negative cases at rates 3 to 5 times that of the random selection method (P < 0.01). The sensitivity for all false-negative cases was 35% and was 52% for false-negative cases at the level of low grade squamous intraepithelial lesion and higher. CONCLUSIONS: The AutoPap 300 QC System provides the potential for a marked increase in the number of false-negative cervical cytology cases that can be detected on QC rescreening. A significant reduction in laboratory false-negative rates can be expected if this device is utilized in routine practice.

Automation↗
PubMed1997

The AutoPap 300 QC System multicenter clinical trials for use in quality control rescreening of cervical smears: II. Prospective and archival sensitivity studies.

BACKGROUND: The AutoPap 300 QC System is an automated device for the analysis of conventional cervical cytology slides. The AutoPap selects an enriched population of cases for quality control review. These studies evaluated the overall sensitivity of AutoPap device to a wider variety of cytologic abnormalities than could be analyzed in the prospective portion of the clinical trials. METHODS: At five clinical trial sites, positive cases were selected from: 1) archives (historic sensitivity study) or 2) current positive cases (current archive sensitivity study). Cases were analyzed by the AutoPap System along with matched negative cases and stratified into detection deciles by instrument score for each diagnostic category. RESULTS: For the historic sensitivity study, the percentages of cases present within the top 10% instrument scores were as follows: atypical glandular cells of undetermined significance (AGUS): 33.7% (n = 243); low grade squamous intraepithelial lesion (LSIL): 57% (n = 412); high grade squamous intraepithelial lesion (HSIL): 81.6% (n = 385); and carcinoma: 77.7% (n = 139). For the current archive sensitivity study, the percentage of cases within the top 10% of instrument scores were as follows: atypical squamous cells of undetermined significance (ASCUS): 37.6% (n = 205); AGUS: 27.3% (n = 22); LSIL: 53.7% (n = 410); HSIL: 80.7% (n = 202); and carcinoma: 62.5% (n = 8). Sensitivities at higher percentiles proportionately were greater. CONCLUSIONS: The AutoPap 300 QC System is detection sensitive, particularly at the level of LSIL and greater. When coupled with the data from the prospective intended use study, these results provided confirmation of the instrument's ability to enhance the false-negative detection rate significantly when compared with a 10% random case selection process. In addition, high sensitivities to all categories of abnormality suggest the possibility of using such instrumentation as a primary screening device for cervical cytology.

Automation↗
PubMed1997

Subtype-specific desensitization of human endothelin ETA and ETB receptors reflects differential receptor phosphorylation.

Endothelins regulate blood pressure in mammals through G protein-coupled receptors. Two receptor subtypes, ETA and ETB, exist which differ by their agonist profiles. Here we show subtype-specific differences in the inactivation of these endothelin receptors. Using a modified inositol phosphate accumulation assay, we found that stimulation of ETA by endothelin-1 results in sustained activation of the subtype, retaining >30% of its initial activity even 20 min after agonist administration, whereas the ETB rapidly deactivated after agonist stimulation, losing >80% of its initial activity within 5 min after endothelin application. The discrepancy in receptor inactivation is reflected by subtype-specific differences in receptor phosphorylation. Whereas ETA failed to undergo ligand-induced phosphorylation, the ETB was rapidly phosphorylated in response to agonist stimulation. By contrast, the kinetics of ligand-induced internalization were essentially identical for the receptor subtypes, suggesting endothelin receptor internalization being independent of ligand-induced receptor phosphorylation. Interestingly, a strong correlation was observed between the time course of ETA inactivation and ETA internalization. Therefore, our data suggest a subtype-specific inactivation of human endothelin receptors: fast receptor phosphorylation in the case of ETB and slow receptor internalization in the case of ETA. Subtype-specific modulation of endothelin receptors may account for the short-term hypotensive effects of endothelins via rapidly down-regulating ETB receptors and the long-lasting hypertensive effects due to sustained ETA activation.

Animals↗
PubMed1997

Application of [(125)I]-[Tyr8]-substance P prepared by the chloramine-T method to receptor-binding experiments after subsequent reduction with mercaptoethanol and purification by reversed-phase liquid chromatography.

Radiolabeling of [Tyr8]-substance P ([Tyr8]-SP) with the (125)I-isotope was performed by use of the chloramine-T technique. The primary formed radiolabeled product, having been quantitatively converted to the corresponding sulfoxide yielding [(125)I]-[Tyr8]-(Met11-->O)-SP completely lacked any binding to proteins rich in SP receptor populations. However, after reductive treatment with mercaptoethanol for about 2 h, a complete reconstitution of the Met11 thioether structure was observed. The reduced peptide, consisting of [(125)I]-[Tyr8]-(Met11)-SP was separated from its by-products by reversed-phase high-performance liquid chromatography on octadecylsilyl silica gel with 100 mM triethyl ammonium formate buffer containing 22% acetonitrile (pH 2.2). The labeled SP derivative prepared by this two-step synthesis was obtained in 73% overall yield related to the [Tyr8]-SP starting material and exhibited a specific activity of 1.9-10(6) Ci/M. In contrast to [(125)I]-[Tyr8]-->(Met11-->O)-SP, satisfactory receptor-binding was now observed with the [(125)I]-->[Tyr8]-(Met11)-SP derivative.

Animals↗
PubMed1997

Molecular forms of somatostatin-like immunoreactivity in the cerebrospinal fluid of patients with senile dementia of the Alzheimer type.

In the cerebrospinal fluid (CSF) of 53 patients with senile dementia of the Alzheimer type (SDAT) and 12 elderly controls, we measured somatostatin (SLI) and its molecular forms: high-molecular weight form (HMV-SST), somatostatin-14 (SST-14), somatostatin-25/28 (SST-28/25), and des-ala-somatostatin (des-ala-SST) using high pressure liquid chromatography (HPLC) and a radioimmunoassay. In SDAT, SLI was significantly decreased (p < 0.05) and correlated with dementia scores (r = -0.65, p < 0.05). HPLC separation showed a marked heterogeneity of SLI in the CSF with a preponderance of SST-14 and SST-25/28. The significant loss of SST-14 (p < 0.05) in SDAT was found to be correlated with dementia scores (r = 0.65). Moreover, qualitative and quantitative changes in the molecular pattern of SLI in SDAT indicated dysregulated synthesis and/or processing of somatostatin relating to the severity of dementia. The long-term administration of neuroleptics in severe cases of SDAT caused a significant increase of SLI (p < 0.05) and influenced the ratio of HMV-SST/SST-14 and SST25/28/SST-14.

Aged↗
PubMed1997

Fine-needle aspiration biopsy of salivary duct carcinoma: report of five cases.

Salivary duct carcinoma is a high grade malignancy which histologically strongly resembles ductal carcinoma of the breast. The findings from five cases of histologically proven salivary duct carcinoma sampled by preoperative fine-needle aspiration (FNA) cytology are presented. Characteristic cytomorphologic features include cohesive clusters and flat sheets of epithelial cells which display a cribriform pattern with eccentrically located, hyperchromatic nuclei, abundant finely granular cytoplasm, and necrosis in the smear background.

Aged↗
PubMed1996

Palmitoylation of endothelin receptor A. Differential modulation of signal transduction activity by post-translational modification.

Post-translational modifications such as phosphorylation and palmitoylation play important roles for the function and regulation of receptors coupled to heterotrimeric guanyl nucleotide-binding proteins. Here we demonstrate that the human endothelin receptor A (ETA) incorporates [3H]palmitate. Mutation of a cluster of five cysteine residues present in the cytoplasmic tail of ETA into serine or alanine residues completely prevented palmitoylation of the receptor. The ligand binding affinity of the non-palmitoylated ETA mutants was essentially unchanged as compared to the palmitoylated wild type ETA suggesting that the replacement of the cysteine residues did not alter the overall structure of the receptor. Furthermore, the ligand-induced stimulation of adenylyl cyclase by the mutant ETA was unaffected by the mutation. In contrast, the mutated non-palmitoylated receptors but not the wild type receptor failed to stimulate phosphatidylinositol hydrolysis by phospholipase C activation upon challenge by endothelin-1. Furthermore, the mutant receptors failed to stimulate the ligand-induced transient increase in the cytoplasmic calcium seen with the wild type ETA. Endothelin-1 induced mitogenic stimuli via the wild type receptors but not through the mutated receptors suggesting an important role for phospholipase C in this signal transduction pathway. The differential regulation of distinct signal transduction pathways by post-translational modification suggests that palmitoylation of the ETA provides a novel mechanism of modulating ETA receptor activity.

Amino Acid Sequence↗
PubMed1996

Human papillomavirus detection by hybrid capture in paired cervicovaginal lavage and cervical biopsy specimens.

Infection of the uterine cervix with human papillomavirus (HPV) is associated with dysplastic lesions that may progress to malignancy. Certain HPV types are associated with higher risk of cervical cancer than other genital HPVs. The goal of this study was to determine if cells obtained by cervicovaginal lavage contain similar HPV types as paired cervical biopsy in women referred because of abnormal cervical cytology. Thirty-four paired lavage and biopsy samples were analyzed for HPV DNA by hybrid capture, using "low risk" (HPV types 6. 11, and related types and "high risk" group (HPV types 16, 18, and related types) HPV. HPV was detected in 24 lavage samples and 18 biopsies. High risk types were predominant. In 14 of 18 HPV-positive biopsies, the paired lavage was also positive for the same HPV group. Four biopsies were HPV-positive at low levels, and the paired lavage was HPV-negative. The mean viral copy numbers of the biopsies from patients with positive and negative lavage samples were 2.7 and 0.1, respectively (P = .02). Ten low level HPV infections were detected by lavage that were not detected by biopsy. HPV detection by hybrid capture in cells obtained by cervicovaginal lavage reflects the results of HPV testing in cervical biopsies.

Cell Biology↗
PubMed1996

Somatostatin, its molecular forms and monoaminergic transmitter metabolites in Binswanger's disease. Neurochemical-neuropathological considerations.

Cerebrospinal fluid (CSF) concentrations of somatostatin-like immunoreactivity (SLI), high molecular weight form somatostatin (HMV-SST), somatostatin-25/28 (SST-25/28), somatostatin-14 (SST-14), Des-ala-somatostatin (Des-ala-SST), homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5-HIAA) were measured in 21 patients with Binswanger's dementia (BD). Patients were classed into three stages of intellectual deterioration according to the Global deterioration scale (GDS). Levels of SLI were significantly decreased in patients suffering from BD, compared to a control group (19.7 +/- 11.6 fmol/ml vs. 30.5 +/- 8.6 fmol/ml, P < 0.01). There was no correlation with dementia scores (r = 0.34, P = 0.51). The observed qualitative and quantitative changes in the molecular pattern of SLI suggest that occurrence of a dysregulated posttranslational processing in patients with BD. Whereas 5-HIAA levels were not significantly changed in patients with BD, HVA was significantly increased in mild to moderate dementia (GDS 2-4) and significantly decreased in severe cases (GDS 7) (224.3 +/- 69.9 nmol/ml vs. 364.9 +/- 103.8 nmol/ml, P < 0.01); this correlated with dementia scores (r = -0.59, P < 0.01). The existence of significant correlations between SLI, 5-HIAA and HVA in BD point to a heterogeneous and generalized neurochemical process affecting several transmitter systems and functions.

Aged↗
PubMed1996

Somatostatin-like immunoreactivity, its molecular forms and monoaminergic metabolites in aged and demented patients with Parkinson's disease--effect of L-Dopa.

There is some evidence that Parkinson's disease (PD) seems to be a heterogenous and generalized brain disorder reflecting a degeneration of multiple neuronal networks, including somatostatinergic neurons. Somatostatin-like immunoreactivity (SLI) and its molecular forms, high molecular weight form (HMV-SST), somatostatin-14 (SST-14), somatostatin-25/28 (SST-25/28) and Des-ala-somatostatin (Des-ala-SST), as well as homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5-HIAA) were estimated using HPLC and radioimmunoassay in the cerebrospinal fluid (CSF) of 35 aged parkinsonian patients with different stages of intellectual deterioration. The influence of L-dopa-treatment on these neurochemical parameters was evaluated. Without a correlation with dementia scores (p = 0.11), SLI was significantly reduced in PD in comparison to the control group (p < 0.05). The reduction was related to the progression of the disease. Correlations between SLI, HVA and 5-HIAA indicate a heterogenous brain disorder in PD with alterations of several transmitter systems and functions. Complex qualitative and quantitative changes in the molecular pattern of SLI are compatible with a dysregulated synthesis and/or posttranslational processing. L-dopa-treatment was associated with a significant increase of HVA (p < 0.05) and HMV-SST (p < 0.05) and a slight, but insignificant increase of SLI (p = 0.11).

Aged↗
PubMed1996

Fine-needle aspiration cytology of lymphoid lesions of the salivary gland: a review of 35 cases.

OBJECTIVE: The aim of this study was to evaluate the effectiveness of fine-needle aspiration (FNA) in the diagnosis of primary lymphoid processes of the salivary gland. DESIGN: A retrospective study. METHOD: Between 1987 and 1994, 35 patients who underwent fine-needle aspiration (FNA) of the salivary gland had a diagnosis of a primary lymphoid process. Most presented with palpable parotid (28 patients) or submandibular (4 patients) gland masses which prompted a clinical diagnosis of pleomorphic adenoma. FNA was performed with immediate on-site evaluation. Flow cytometric cell-surface-marker analysis was performed in 28 of the 35 cases to determine the clonality of the B-cell proliferations. RESULTS: Sixteen cases of reactive hyperplasia and nine cases of malignant lymphoma diagnosed by FNA were confirmed by subsequent histopathologic examination. Lymphoma was confirmed in six of eight cases diagnosed as suspicious for lymphoma by FNA. Hodgkin's disease was suspected but not confirmed in one case and was diagnosed as a probable Warthin's tumour in another case. In all cases, the FNA diagnosis of either a reactive or malignant lymphoid process was unexpected and influenced the patient's further management. For patients diagnosed with an intraparotid lymph node, surgery could be deferred for a short period with the hope that the lymphadenopathy would spontaneously regress. For patients with a preoperative FNA diagnosis of lymphoma, a more limited biopsy could be performed, thereby reducing the operative risk to the patient and plans to process the tissue according to the institution's lymphoma protocol could be made. CONCLUSION: Clinically, reactive intraparotid lymph nodes and lymphomas present as parotid enlargements that are indistinguishable from pleomorphic adenomas. FNA is the only method of accurately establishing a preoperative diagnosis in these patients.

Adolescent↗
PubMed1994

CEO summit. The new delivery & financing realities. Part III of III.

In cooperation with McManis Associates Inc., Washington, Hospitals & Health Networks recently convened a summit on the integration of financing and delivery in health care. This installment is the third of a three-part series on lessons learned by those on the front lines of integration activity. The session was designed and facilitated by senior associates at McManis. Among the issues summit participants discussed in the second segment: What level of understanding do purchasers have of the factors that differentiate quality in health care services? Can provider-driven integrated delivery systems compete with insurer-driven ones? And what happens when a large integrated delivery system merges with a dominant insurer, as happened in the Philadelphia market? Can that model be successfully replicated in other markets? In this final segment, participants talk about whether providers' deep connections to their communities will add value in a reformed delivery system; how incentives might be aligned among all the players in integrated networks and organizations; how the concept of community focus might be redefined under systems integration; and the process involved in preparing for constant, accelerated change. The second segment concluded with comments about the assets providers and insurers bring to integrated health systems, and whether the merger experience of Graduate Health System and QCC/Independence Blue Cross could be replicated in other markets or not.

Community-Institutional Relations↗