Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “multiple clustering”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 811 records · Page 45Linked to original sources
PubMed1991

AgNOR (nucleolar organizer regions) staining in malignant melanoma.

Nucleolar organizer regions (NORs) are loops of ribosomal DNA seen in nuclei, which are demonstrable as black dots (AgNOR) in tissue sections by silver (Ag) colloid staining. The number of such AgNORs is correlated with cellular activity and is an indicator of the degree of malignancy. In this study, 76 melanocytic lesions were analyzed by AgNOR staining, and the clinical and histopathological characteristics of malignant melanoma and melanocytic nevi were considered. Although the AgNOR counts for melanocytic nevi were significantly different from those in malignant melanoma, an obvious overlap between them was detected. The number of AgNORs in melanocytic nevi per cell was usually 1 or 2. On the other hand, the number of AgNORs per malignant melanoma cell was variable. Morphologically, malignant melanoma cells often showed dispersal of AgNORs throughout the nucleus as well as multiple nucleoli containing clustered AgNORs, whereas melanocytic nevus cells tended to have a regular nucleolus with tightly clustered AgNORs. The correlation between AgNOR count and pathological staging was uncertain, but a slight correlation between AgNOR count and thickness of the primary lesion was obtained. However, the AgNOR count in malignant melanoma was not a prognostic factor for the disease. Therefore, the AgNOR method is difficult to use for differential diagnosis between benign pigmented lesions and malignant melanoma. Nonetheless, an AgNOR count of more than two per cell favors a diagnosis of malignant melanoma.

Adult↗
PubMed2004

Increased risk of skin cancer associated with the presence of epidermodysplasia verruciformis human papillomavirus types in normal skin.

BACKGROUND: Human papillomaviruses (HPVs) are found in normal skin and in benign and malignant skin conditions. Epidermodysplasia verruciformis (EV) HPV types are those most plausibly linked to the development of squamous cell carcinomas of the skin. OBJECTIVES: To assess the risk of nonmelanoma skin cancer (NMSC) associated with the presence of EV HPV in normal skin in immunocompetent (IC) individuals and renal transplant recipients (RTRs). METHODS: Using a degenerate and nested polymerase chain reaction technique, HPV DNA was sought in 124 normal skin samples from sun-exposed and nonsun-exposed sites, from 39 IC individuals and 38 RTRs, both with and without NMSC. Data were analysed using the Mantel-Haenszel test and by logistic regression analysis. RESULTS: HPV DNA was detected in 58/67 (87%) and 20/57 (35%) samples from renal transplant and IC patients, respectively. There was no difference in either the prevalence or spectrum of HPV types found in sun-exposed and nonsun-exposed normal skin. However, there was significant association between NMSC and the presence of EV HPV DNA. Multivariate analysis provided an odds ratio of 6.41 (95% confidence interval 1.79-22.9) for the association of EV HPV DNA in normal skin (irrespective of site) and NMSC status, even after stratifying for patient group and adjusting for the clustering effect of multiple sampling. Conversely, there was no association between skin cancer status and the presence of cutaneous or mucosal HPV types in either sun-exposed or nonsun-exposed skin. CONCLUSIONS: HPV DNA is widespread in normal adult skin, particularly in transplant patients. In our study, the presence of EV but not cutaneous HPV DNA in normal skin was significantly associated with NMSC status and may prove to be of predictive value for skin cancer risk. These data provide reason to focus on EV HPV types as causal agents in skin cancer.

Adult↗
PubMed1981

Replication of ribosomal DNA in Xenopus laevis.

The study of the localization of the replication origins of rDNA in Xenopus laevis has been approached by two different methods. 1. The DNA of X. laevis larvae was fractionated by CsCl gradient centrifugation in bulk and ribosomal DNA and examined in the electron microscope. In bulk DNA, clusters of microbubbles, which are related with the origins of replication, appear to be spaced along the DNA molecules at intervals comparable with the size of the 'average' replicon of X. laevis. In ribosomal DNA, the distance between adjacent clusters is much shorter and corresponds to the size of the rDNA repeating unit. When ribosomal DNA was submitted to digestion with restriction enzymes (Eco RI and HindIII) the microbubbles are observed in the non-transcribed spacer-containing fragment. 2. Cultured cells of X. laevis were synchronized by mitotic selection and incubated with 5-fluoro-2-deoxyuridine for a time longer than the G1 phase. This treatment synchronizes the replicons and allows them to start replicating very slowly. It was thus possible to obtain a preferential labelling of the regions containing the origins. The analysis by gel electrophoresis of the Eco Ri-digested rDNA showed that the radioactivity was preferentially incorporated in the fragments which contain the non-transcribed spacer. The results of these two approaches indicate that the rRNA gene cluster consists of multiple units of replication, possibly one per gene unit. Furthermore they show that the origins of replication are localized into the non-transcribed spacer.

Animals↗
PubMed2006

Role of glycoprotein Ibalpha in phagocytosis of platelets by macrophages.

BACKGROUND: Platelet (PLT) storage at 0 to 4 degrees C suppresses bacterial multiplication, but induces clusters of glycoprotein (GP) Ibalpha that trigger their phagocytosis by macrophages and reduce their survival after transfusion. A method was sought that detects cold-induced changes in GPIbalpha involved in phagocytosis. STUDY DESIGN AND METHODS: Human PLTs were isolated and stored for up to 48 hours at 0 degrees C. Binding of a phycoerythrin (PE)-labeled antibody directed against amino acids (AA) 1-35 on GPIbalpha (AN51-PE) was compared with phagocytosis of PLTs by matured monocytic THP-1 cells, analyzed by fluorescence-activated cell sorting. RESULTS: Freshly isolated PLTs were detected as a single population of AN51-PE-positive particles and showed less than 5 percent phagocytosis. Cold storage led to a decrease in AN51-PE binding and an increase in phagocytosis. N-Acetylglucosamine, known to interfere with macrophage recognition of GPIbalpha clusters, restored normal AN51-PE binding to cold-stored PLTs and suppressed phagocytosis. CONCLUSIONS: It is concluded that binding of an antibody against AA 1-35 on GPIbalpha reflects changes in GPIbalpha that make PLTs targets for phagocytosis by macrophages.

Acetylglucosamine↗
PubMed2006

Sleep and the metabolic syndrome.

The metabolic syndrome represents a clustering of several interrelated risk factors of metabolic origin that are thought to increase cardiovascular risk. It is still uncertain whether this clustering results from multiple underlying risk factors or whether it has a single cause. One metabolic abnormality that may underlie several clinical characteristics of the metabolic syndrome is insulin resistance. This review discusses the evidence that sleep disturbances (obstructive sleep apnoea, sleep deprivation and shift work) may independently lead to the development of both insulin resistance and individual clinical components of the metabolic syndrome. The converse may also be true, in that metabolic abnormalities associated with the metabolic syndrome and insulin resistance may potentially exacerbate sleep disorders. The notion that sleep disturbances exert detrimental metabolic effects may help explain the increasing prevalence of the metabolic syndrome and insulin resistance in the general population and may have important implications for population-based approaches to combat the increasing epidemic of metabolic and cardiovascular disease.

Animals↗
PubMed1996

Population structure of Streptococcus agalactiae reveals an association between specific evolutionary lineages and putative virulence factors but not disease.

To evaluate the genetic diversity and relationships in a collection of 85 Danish strains of Streptococcus agalactiae (group B streptococcus) we have performed restriction fragment length polymorphism analysis on EcoRI- and MspI-digested whole-cell DNA using as probes rRNA, DNA fragments representing the genes encoding hyaluronidase, C5a-peptidase, alpha-antigen, and beta-antigen as well as two randomly selected genomic DNA fragments for which the coding potential is unknown. In addition, we have assayed for expression of hyaluronidase activity and beta-antigen. Combined analyses of our data and those previously obtained by multilocus enzyme electrophoresis and serotyping revealed a population separating into six major lineages that correlate with individual serotypes. The significant linkage disequilibrium of alleles indicates that the S. agalactiae population examined is predominantly clonal. Notably, strains expressing the serotype III capsule divide into two distant evolutionary lineages, of which one lacks expression of hyaluronidase activity. Six North American isolates of serotype III clustered together with multiple Danish serotype III strains, showing that the combinations of characters on which the phylogenetic tree was based are conserved worldwide. Occurrence of beta-antigen correlated with a specific version of the alpha-antigen gene and was exclusively associated with a single major phylogenetic lineage. Comparisons with the clinical history of the strains revealed no evidence of differences in pathogenic potential among the six major genetic divisions.

Adhesins, Bacterial↗
PubMed2006

Type III pilus of corynebacteria: Pilus length is determined by the level of its major pilin subunit.

Multiple pilus gene clusters have been identified in several gram-positive bacterial genomes sequenced to date, including the Actinomycetales, clostridia, streptococci, and corynebacteria. The genome of Corynebacterium diphtheriae contains three pilus gene clusters, two of which have been previously characterized. Here, we report the characterization of the third pilus encoded by the spaHIG cluster. By using electron microscopy and biochemical analysis, we demonstrate that SpaH forms the pilus shaft, while SpaI decorates the structure and SpaG is largely located at the pilus tip. The assembly of the SpaHIG pilus requires a specific sortase located within the spaHIG pilus gene cluster. Deletion of genes specific for the synthesis and polymerization of the other two pilus types does not affect the SpaHIG pilus. Moreover, SpaH but not SpaI or SpaG is essential for the formation of the filament. When expressed under the control of an inducible promoter, the amount of the SpaH pilin regulates pilus length; no pili are assembled from an SpaH precursor that has an alanine in place of the conserved lysine of the SpaH pilin motif. Thus, the spaHIG pilus gene cluster encodes a pilus structure that is independently assembled and antigenically distinct from other pili of C. diphtheriae. We incorporate these findings in a model of sortase-mediated pilus assembly that may be applicable to many gram-positive pathogens.

Bacterial Proteins↗
PubMed1992

Isolation and analysis of a novel inducible pectate lyase gene from the phytopathogenic fungus Fusarium solani f. sp. pisi (Nectria haematococca, mating population VI).

A pectate lyase produced by Fusarium solani f. sp. pisi (Nectria haematococca, mating population VI) was previously shown to be essential for host infection (M. S. Crawford and P. E. Kolattukudy, Arch. Biochem. Biophys. 258:196-205, 1987). Pectate lyase genes have not been cloned from any phytopathogenic fungi. A gene, designated pelA, encoding an inducible pectate lyase was isolated from F. solani f. sp. pisi. A probe was synthesized by polymerase chain reaction with oligonucleotide primers based on the known amino acid sequences of two regions of the mature protein and first-strand cDNA as template. Both cDNA and the gene were isolated and sequenced. That the cloned cDNA represents the previously purified pectate lyase is shown by the complete match of the sequences of the N-terminal 38 amino acid residues and the 20 amino acid residues of an internal peptide with the sequence deduced from the cDNA sequence. This lyase sequence shows little homology to those of other pectolytic enzymes. The pelA gene shows standard characteristics with respect to promoter, intron, and polyadenylation sequences. As determined by primer extension and nuclease S1 analysis of the origin of the transcription, there are multiple initiation sites clustered in a region of 12 nucleotides located about 55 bp upstream of the start codon. Northern (RNA) blot analysis showed a single band of mRNA at about 1 kb. The pelA gene mRNA was detected only when F. solani f. sp. pisi was grown with pectin, and there was no detectable transcript accumulation when the fungus was grown with glucose as the sole carbon source. When both carbon sources were present, the pelA gene was transcribed only after glucose was completely depleted, indicating carbon catabolite repression. Moreover, the levels of transcription decreased rapidly prior to maximal enzyme accumulation, suggesting a mechanism of self catabolite repression.

Amino Acid Sequence↗
PubMed1994

Metabolism of naphthalene, fluorene, and phenanthrene: preliminary characterization of a cloned gene cluster from Pseudomonas putida NCIB 9816.

A modified cloning procedure was used to obtain large DNA insertions (20 to 30 kb) from Pseudomonas putida NCIB 9816 that expressed polycyclic aromatic hydrocarbon (PAH) transformation activity in Escherichia coli HB101. Four subclones (16 [in both orientations], 12, and 8.5 kb in size) were constructed from the initial clones. Naphthalene, fluorene, and phenanthrene transformations were investigated in these eight NCIB 9816 clones by a simple agar plate assay method, which was developed to detect and identify potential PAH metabolites. Results indicated that the necessary genes encoding the initial ring fission of the three PAHs in E. coli cells are located in an 8.5-kb EcoRI-XhoI portion, but the lower-pathway genes are not present in a 38-kb neighborhood region. These NCIB 9816 clones could transform naphthalene and phenanthrene to salicylic acid and 1-hydroxy-2-naphthoic acid, respectively. With the same clones, fluorene was degraded to 9-hydroxyfluorene, 9-fluorenone, and two unidentified compounds. Genetic similarity between the NAH7 upper-pathway genes and the cloned NCIB 9816 genes was confirmed by Southern blot DNA-DNA hybridization. In spite of this genetic similarity, the abilities of the two clusters to transform multiple PAHs were different. Under our experimental conditions, only the metabolites from naphthalene transformation by the NAH7 clone (pE317) were detected, whereas the NCIB 9816 clones produced metabolites from all three PAHs.

Biotransformation↗
PubMed2006

Assembly of distinct pilus structures on the surface of Corynebacterium diphtheriae.

Different surface organelles contribute to specific interactions of a pathogen with host tissues or infectious partners. Multiple pilus gene clusters potentially encoding different surface structures have been identified in several gram-positive bacterial genomes sequenced to date, including actinomycetales, clostridia, corynebacteria, and streptococci. Corynebacterium diphtheriae has been shown to assemble a pilus structure, with sortase SrtA essential for the assembly of a major subunit SpaA and two minor proteins, SpaB and SpaC. We report here the characterization of a second pilus consisting of SpaD, SpaE, and SpaF, of which SpaD and SpaE form the pilus shaft and SpaF may be located at the pilus tip. The structure of the SpaDEF pilus contains no SpaABC pilins as detected by immunoelectron microscopy. Neither deletion of spaA nor sortase srtA abolishes SpaDEF pilus formation. The assembly of the SpaDEF pilus requires specific sortases located within the SpaDEF pilus gene cluster. Although either sortase SrtB or SrtC is sufficient to polymerize SpaDF, the incorporation of SpaE into the SpaD pili requires sortase SrtB. In addition, an alanine in place of the lysine of the SpaD pilin motif abrogates pilus polymerization. Thus, SpaD, SpaE, and SpaF constitute a different pilus structure that is independently assembled and morphologically distinct from the SpaABC pili and possibly other pili of C. diphtheriae.

ATP-Binding Cassette Transporters↗
PubMed1990

Cell type-specific protein-DNA interactions in the human zeta-globin upstream promoter region: displacement of Sp1 by the erythroid cell-specific factor NF-E1.

The protein-DNA interactions of the upstream promoter region of the human embryonic zeta-globin gene in nuclear extracts of erythroid K562 cells and nonerythroid HeLa cells were analyzed by DNase I footprinting, gel mobility shift assay, methylation interference, and oligonucleotide competition experiments. There are mainly two clusters of nuclear factor-binding sites in the zeta promoter. The proximal cluster spans the DNA sequence from -110 to -60 and consists of binding sites for CP2, Sp1, and NF-E1. NF-E1 binding is K562 specific, whereas CP2 binding is common to both types of cells. Overlapping the NF-E1- and CP2-binding sites is a hidden Sp1-binding site or CAC box, as demonstrated by binding studies of affinity-purified Sp1. In the distal promoter region at -250 to -220, another NF-E1-binding site overlaps a CAC box or Sp1-binding site. Extract-mixing experiments demonstrated that the higher affinity of NF-E1 binding excluded the binding of Sp1 in the K562 extract. NF-E1 factors could also displace prebound Sp1 molecules. Between the two clusters of multiple-factor-binding sites are sequences recognized by other factors, including zeta-globin factors 1 and 2, that are present in both HeLa and K562 extracts. We discuss the cell type-specific, competitive binding of multiple nuclear factors in terms of functional implications in transcriptional regulation of the zeta-globin gene.

Base Sequence↗
PubMed2004

Recruitment of the nucleolar remodeling complex NoRC establishes ribosomal DNA silencing in chromatin.

The rRNA gene cluster consists of multiple transcription units. Half of these are active, while the other half are transcriptionally inactive. Previously, in vivo studies have demonstrated that silencing of ribosomal DNA (rDNA) is mediated by the chromatin remodeling NoRC (nucleolar remodeling complex). To explore the mechanisms underlying NoRC-directed silencing of rDNA transcription, we investigated the effect of recombinant NoRC on RNA polymerase I transcription on reconstituted chromatin templates. We show that NoRC interacts with the transcription terminator factor (TTF-I), and this interaction is required both for the binding of TTF-I to its promoter-proximal target site and for the recruitment of NoRC to the promoter. After association with the rDNA promoter, NoRC alters the position of the promoter-bound nucleosome, thereby repressing RNA polymerase I transcription. This NoRC-directed rDNA repression requires the N terminus of histone H4. Repression is effective before preinitiation complex formation and as such is unable to exert an effect upon activated rDNA genes. Furthermore, the early steps of rDNA repression do not depend on DNA and histone modifications. These results reveal an important role for TTF-I in recruiting NoRC to rDNA and an active role for NoRC in the establishment of rDNA silencing.

Acetylation↗
PubMed2026

Draft genome sequence of Bacillus thuringiensis GIFSPR-111, a putative probiotic isolated from shrimp farm soil in Bangladesh.

Bacillus thuringiensis GIFSPR-111, isolated from shrimp farm soil in Shamnagar, Bangladesh, inhibits Vibrio parahaemolyticus, the causative agent of acute hepatopancreatic necrosis disease in shrimp and exhibits probiotic potential. The 5,608,442 bp draft genome contains multiple biosynthetic gene clusters, with potential to synthesize diverse bioactive metabolites.

Bacillus thuringiensis↗
PubMedDate not supplied

Thin-layer cytology findings of papillary adenosquamous carcinoma of the cervix. Report of a case with histologic correlation and molecular analysis.

BACKGROUND: Papillary squamotransitional cell carcinoma of the lower female genital tract is a rare form of malignancy with distinctive histologic features. To my knowledge, its adenosquamous counterpart has not been reported before. CASE: A 69-year-old woman presented with postmenopausal bleeding due to a bulky cervical tumor. The thin-layer cytology preparation (Autocyte, TriPath Imaging, Burlington, North Carolina, U.S.A.) was of high cellularity and contained multiple small, papillary clusters of basaloid to columnar cells. Discernible fibrovascular cores were noted. In the background were many loosely dispersed, bland-looking columnar cells as well as cells of high grade squamous intraepithelial lesion (CIN 3). On a diligent search, scattered adenocarcinoma cells containing distinct intracytoplasmic vacuoles were found. There were a small number of dyskaryotic squamous cells with bizarre shapes, assuming "tadpolelike" morphology. Histologic examination of the tumor biopsy showed a papillary tumor with central fibrovascular cores covered with mitotically active, poorly differentiated squamous cells that imperceptibly merged with abortive glandular structures. Molecular analysis using the polymerase chain reaction and restriction fragment length polymorphism revealed the presence of human papillomavirus type 16 DNA in the liquid-based cytology sample. CONCLUSION: Papillary adenosquamous carcinoma of the cervix shares some morphologic and even histogenetic overlaps with papillary squamotransitional cell carcinoma and, to a lesser extent, conventional endocervical adenocarcinoma. It may pose certain diagnostic dilemmas for the unwary. In view of its distinctive cytohistologic features, this tumor warrants greater recognition in gynecologic pathology.

Aged↗
PubMed2006

A graphical model approach to automated classification of protein subcellular location patterns in multi-cell images.

BACKGROUND: Knowledge of the subcellular location of a protein is critical to understanding how that protein works in a cell. This location is frequently determined by the interpretation of fluorescence microscope images. In recent years, automated systems have been developed for consistent and objective interpretation of such images so that the protein pattern in a single cell can be assigned to a known location category. While these systems perform with nearly perfect accuracy for single cell images of all major subcellular structures, their ability to distinguish subpatterns of an organelle (such as two Golgi proteins) is not perfect. Our goal in the work described here was to improve the ability of an automated system to decide which of two similar patterns is present in a field of cells by considering more than one cell at a time. Since cells displaying the same location pattern are often clustered together, considering multiple cells may be expected to improve discrimination between similar patterns. RESULTS: We describe how to take advantage of information on experimental conditions to construct a graphical representation for multiple cells in a field. Assuming that a field is composed of a small number of classes, the classification accuracy can be improved by allowing the computed probability of each pattern for each cell to be influenced by the probabilities of its neighboring cells in the model. We describe a novel way to allow this influence to occur, in which we adjust the prior probabilities of each class to reflect the patterns that are present. When this graphical model approach is used on synthetic multi-cell images in which the true class of each cell is known, we observe that the ability to distinguish similar classes is improved without suffering any degradation in ability to distinguish dissimilar classes. The computational complexity of the method is sufficiently low that improved assignments of classes can be obtained for fields of twelve cells in under 0.04 second on a 1600 megahertz processor. CONCLUSION: We demonstrate that graphical models can be used to improve the accuracy of classification of subcellular patterns in multi-cell fluorescence microscope images. We also describe a novel algorithm for inferring classes from a graphical model. The performance and speed suggest that the method will be particularly valuable for analysis of images from high-throughput microscopy. We also anticipate that it will be useful for analyzing the mixtures of cell types typically present in images of tissues. Lastly, we anticipate that the method can be generalized to other problems.

Algorithms↗
PubMedDate not supplied

Symptoms experience: a concept analysis.

PURPOSE/OBJECTIVES: To provide a clearly constructed definition of the concept of symptoms experience. DATA SOURCES: Articles and book chapters. DATA SYNTHESIS: Symptoms experience has not been explored previously. Several approaches to the concept of symptoms have been addressed, including symptom occurrence, symptom distress, and unpleasant symptoms. Limitations of these approaches may include the lack of focus on symptoms as occurring concurrently or in clusters or the multiplicative nature of symptoms. In addition, situational and existential meaning often is not explored. CONCLUSIONS: Symptoms experience is the perception of the frequency, intensity, distress, and meaning of symptoms as they are produced and expressed. Symptoms are multiplicative in nature and may act as catalysts for the occurrence of other symptoms. Antecedents to the symptoms experience include demographic, disease, and individual factors. Consequences include the impact on mood state, psychological status, functional status, quality of life, disease progression, and survival. IMPLICATIONS FOR NURSING: Evaluation of symptoms in patients with cancer should include a meaning-centered approach, in which symptoms are evaluated not only for occurrence characteristics and perceived distress but also for the meaning of the symptoms experience to individuals.

Adult↗
PubMed1991

Dynamics of pulsatile prolactin release during the postpartum lactational period.

To investigate the pulsatile nature of PRL release in the physiologically hyperprolactinemic postpartum period, we sampled blood at 10-min intervals for 24 h in each of 6 healthy lactating women at both 3 weeks and 3 months postpartum. The subsequent immunoactive PRL time series were subjected to episodic peak detection (Cluster analysis) and multiple parameter deconvolution analysis. The 24-h mean serum PRL concentrations were significantly higher at 3 weeks than at 3 months postpartum; viz. 113 +/- 12 vs. 66 +/- 15 micrograms/L (P = 0.003). Assessment of episodic PRL pulsatility revealed significantly higher maximal PRL peak heights (296 +/- 63 vs. 141 +/- 44 micrograms/L), fractional peak heights (863 +/- 150 vs. 374 +/- 58%), incremental peak amplitudes (250 +/- 60 vs. 96 +/- 3 micrograms/L), and peak areas (13 +/- 3 vs. 4 +/- 1 mg/L.min) in the earlier postpartum period. In contrast, PRL peak frequencies and interpulse intervals were not different in the early and late postpartum sessions. Deconvolution analysis revealed that the mean mass of PRL secretory bursts was significantly greater at 3 weeks (182 +/- 4.1 micrograms/L) than 3 months (15 +/- 1.6 micrograms/L). There were no changes in the calculated half-life of endogenous PRL viz. 29 +/- 2.5 min (3 weeks) vs. 26 +/- 3.0 min (3 months). We conclude that physiological postpartum hyperprolactinemia is achieved by selectively altering the endogenous secretory rate in each PRL release episode, with no change in the number of bursts of PRL discharged or the PRL half-life.

Adult↗
PubMed2002

Pulsatile insulin secretion by human pancreatic islets.

Insulin is secreted in discrete bursts. These pulses are also present when individual or groups of islets are perifused. Interpretation of the measured frequency and magnitude of pulsatile hormone secretion requires an examination of the sensitivity and specificity of the methods for pulse detection and validation of these for the experimental apparatus and hormone assay in which they are applied. In the present study we achieve these aims for a perfusion method for measurement of pulsatile insulin release by human islets. A deconvolution technique previously developed for measurement of pulsatile hormone secretion in vivo was specifically validated for in vitro pulse detection in the present study. Deconvolution analysis reliably (>90%) detected insulin pulses with an amplitude 20% or more above baseline and recovered quantitatively the insulin secretion profile, insulin secretion rate, and insulin pulse mass from single as well as multiple perifused islets. Cluster analysis was less sensitive, but was able to detect most (>80%) pulses with an amplitude of 40% or more above baseline. With this limitation, cluster analysis is potentially useful for groups, but not single perifused human islets. Analysis of single human islets showed that enhanced insulin secretion by increased glucose concentrations in the perfusate is achieved by enhancing insulin pulse mass with no change in pulse frequency. Perfused single or groups of human islets exhibited an interpulse interval ( approximately 6-8 min) comparable to that observed in humans in vivo. Dynamic in vitro perifusion should facilitate studies of the mechanisms driving pulsatile insulin secretion.

Cluster Analysis↗