Search PubMed⌕ Search

Biomedical subjects

M Kremer

Publications and source records attributed to M Kremer.

At least 37 records · Page 2Linked to original sources
PubMed2000

Connexin43 null mice reveal that astrocytes express multiple connexins.

The gap junction protein connexin43 (Cx43) is the primary component of intercellular channels in cardiac tissue and in astrocytes, the most abundant type of glial cells in the brain. Mice in which the gene for Cx43 is deleted by homologous recombination die at birth, due to profound hypertrophy of the ventricular outflow tract and stenosis of the pulmonary artery. Despite this significant cardiovascular abnormality, brains of connexin43 null [Cx43 (-/-)] animals are shown to be macroscopically normal and to display a pattern of cortical lamination that is not detectably different from wildtype siblings. Presence of Cx40 and Cx45 in brains and astrocytes cultured from both Cx43 (-/-) mice and wildtype littermates was confirmed by RT-PCR, Northern blot analyses and by immunostaining; Cx46 was detected by RT-PCR and Northern blot analyses. Presence of Cx26 in astrocyte cultures was indicated by RT-PCR and by Western blot analysis, although we were unable to resolve whether it was contributed by contaminating cells; Cx30 mRNA was detected by Northern blot in long term (2 weeks) but not fresh cultures of astrocytes. These studies thus reveal that astrocyte gap junctions may be formed of multiple connexins. Presumably, the metabolic and ionic coupling provided by these diverse gap junction types may functionally compensate for the absence of the major astrocyte gap junction protein in Cx43 (-/-) mice, providing whatever intercellular signaling is necessary for brain development and cortical lamination.

Animals↗
PubMed2000

PCR analysis of IgH-gene rearrangements in small lymphoid infiltrates microdissected from sections of paraffin-embedded bone marrow biopsy specimens.

The differentiation of benign lymphoid infiltrates from nodular infiltrates of B-cell lymphoma is difficult in bone marrow (BM) biopsy specimens taken from patients with non-Hodgkin's lymphoma (NHL). We investigated whether the determination of clonality by polymerase chain reaction (PCR) analysis of the immunoglobulin heavy chain (IgH) genes could be of help for the distinction of benign and malignant lymphoid infiltrates. BM biopsy specimens of 28 patients were studied, comparing PCR of entire bone marrow sections with microdissected nodular lymphoid infiltrates. Patients were divided into 4 groups according to morphologic criteria: group 1 (n = 12), positive for B-NHL infiltration; group 2 (n = 5), suspicious for infiltration by known B-NHL; group 3 (n = 5), morphologically benign infiltrates in patients with B-NHL; group 4 (n = 6), benign lymphoid infiltrates in patients without history of B-NHL. PCR products were analyzed using polyacrylamide gels and a fragment length analysis system (Genescan). PCR of whole sections showed clonal amplification products in all cases of group 1 and 1 case of group 2. PCR analysis from microdissected nodular infiltrates showed the presence of a clonal B-cell population in 5 additional cases of groups 2 and 4. In 3 of these cases, clonal rearrangements of corresponding size were obtained from the primary lymphoma biopsy specimens. None of the cases of group 3 showed evidence of a clonal population with either technique. The results indicate that microdissection of small nodular lymphoid infiltrates from paraffin-BM sections increases the sensitivity of IgH gene rearrangement analysis. To avoid detection of biologically irrelevant clonal populations, comparison of PCR products obtained from the BM and the primary lymphoma biopsy is advisable.

Biopsy↗
PubMed2000

Evaluation of dermal-epidermal skin equivalents ('composite-skin') of human keratinocytes in a collagen-glycosaminoglycan matrix(Integra artificial skin).

Integra artificial skin (Integra LifeSciences Corp., Plainsboro, NJ, USA) is a dermal template consisting of bovine collagen, chondroitin-6-sulphate and a silastic membrane manufactured as Integra. This product has gained widespread use in the clinical treatment of third degree burn wounds and full thickness skin defects of different aetiologies. The product was designed to significantly reduce the time needed to achieve final wound closure in the treatment of major burn wounds, to optimise the sparse autologous donor skin resources and to improve the durable mechanical quality of the skin substitute. The clinical procedure requires two stages. The first step creates a self neodermis, the second creates a self epidermis on the neodermis. However, it is desirable to cover major burn wounds early in a single step by a skin substitute consisting of a dermal equivalent seeded in vitro with autologous keratinocytes ('composite-skin') out of which a full thickness skin develops in vivo.The goal of this experimental study was to develop a method to integrate human keratinocytes in homogeneous distribution and depth into Integra Artificial Skin. The seeded cell-matrix composites were grafted onto athymic mice in order to evaluate their potential to reconstitute a human epidermis in vivo. We were able to demonstrate that the inoculated human keratinocytes reproducibly displayed a homogeneous pattern of distribution, adherence, proliferation and confluence. The cell-matrix composites grafted in this model exhibited good wound adherence, complete healing, minor wound contraction and had the potential to reconstitute an elastic, functional and durable human skin. Histologically we were able to show that the inoculated human keratinocytes in vivo colonised the matrix in a histomorphologically characteristic epidermal pattern (keratomorula, keratinocyte bubbling) and developed a persisting, stratified, keratinising epidermis which immunohistologically proved to be of human origin. These experimental results demonstrate the establishment of an effective cell cultivation process which may be suitable for scale-up production of the epidermal component as large-scale composite-skin grafts. When seeded into Integratrade mark and grafted onto the nude mouse a replacement skin with normal functioning dermal-epidermal components was developed. These results encourage the design of a clinical trial to assess the function of this composite graft in man.

Animals↗
PubMed2000

Accumulation of chromosomal imbalances from intraductal proliferative lesions to adjacent in situ and invasive ductal breast cancer.

Carcinoma of the breast is thought to evolve through a sequential progression from normal to proliferative epithelium and eventually into carcinoma. Here lumpectomy specimens from five patients were studied, selected for the presence of ductal hyperplasia without atypia, atypical ductal hyperplasia, ductal carcinoma in situ, and invasive ductal carcinoma. Laser microdissection of tissue allowed precise sampling and direct correlation of phenotypic and genotypic changes. Analyses of the samples revealed an increasing mean number of chromosomal changes occurring with increasing histologic severity, and for the first time chromosomal abnormalities were demonstrated in ductal hyperplasia without atypia. Chromosomal changes found in each of the four histologic entities included gains on 10q, 12q, 16p, and 20q and loss on 13q. In ductal hyperplasia without atypia, gain on 20q as well as loss on 13q was detected with high frequency (four of five samples). Alterations identified in more than 50% of atypical ductal hyperplasia samples included gains on 3p, 8q, 15q, and 22q and loss on 16q. In ductal carcinoma in situ, gain of DNA on 1q and 17q and loss on 4q were additionally found, and in invasive ductal carcinoma, further gains on 6p, 10q, 11q13, and 17p were identified. The chromosomal alterations occurring in the different histopathologic lesions strongly suggest that these regions harbor tumor suppressor genes or oncogenes significant for the development of ductal carcinoma of the breast.

Breast↗
PubMed2000

Laser capture microdissection: methodical aspects and applications with emphasis on immuno-laser capture microdissection.

Laser capture microdissection (LCM) is an easy, extremely fast and versatile method for the isolation of morphologically defined cell populations from complex primary tissues for molecular analyses. However, the optical resolution is limited due to the use of dried sections without coverslip necessary for tissue capture, and routine stains such as hematoxylin and eosin are sometimes insufficient for precise microdissection, especially in tissues with diffuse intermingling of different cell types and lack of easily discernible architectural features. Therefore, several groups have adapted immunohistochemical staining techniques for LCM. In addition to providing high contrast targets for microdissection, immunostaining allows selection of cells not only according to morphological, but also phenotypical and functional criteria. In order to allow reliable tissue transfer on one hand and preserve the integrity of the target of analysis such as DNA, RNA and proteins on the other hand, immunostaining protocols have to be modified for the purposes of LCM. The following review gives an overview of immuno-LCM and describes some applications, e.g. in the field of hematopathology.

Cell Separation↗
PubMed2000

Epidemiology of single and multiple species of helminth infections among school children in Busia District, Kenya.

OBJECTIVE: To describe the patterns of single and multiple helminth infection in school children from Busia District, Kenya. DESIGN: A cross-sectional school survey using a randomly selected sample, forming part of an evaluation study of an ongoing deworming project. SETTING: Budalangi and Funyula divisions of Busia District, Western Province, Kenya. SUBJECTS: One thousand seven hundred and thirty eight school children aged 8-20 years randomly selected from those enrolled in standards 3-8 in 25 randomly selected primary schools. RESULTS: Overall, 91.7% of children were infected with either hookworm, Ascaris lumbricoides, Trichuris trichiura or Schistosoma mansoni. Infection prevalence of each species varied considerably among schools, being most marked for S. mansoni, where prevalence was highest in lakeshore schools. Children were typically infected with two or more species of helminth. Infection intensity of each geohelminth species was higher in school children infected with multiple species than in school children with single species infections, and intensity increased with the number of concurrent infections. CONCLUSION: Helminth infections are exceptionally common among school children in Busia district, thus confirming the good sense of the school-based approach adopted by the control programme. The study also shows that there is an association between concurrent infection and the intensity of infection, which may have consequences for nutritional and educational status.

Adolescent↗
PubMed1999

Transcription factor YY1 is a vaccinia virus late promoter activator.

Vaccinia virus has a DNA genome, yet replicates in the cytoplasmic compartment of the cell. We previously described the identification of a cellular protein having high affinity for vaccinia virus late promoter DNA. Sequence substitutions in the vaccinia I1L promoter were used to define a 5-nucleotide block at the transcription initiation site as essential for interaction with the protein. Within this sequence is the recognition motif for the nuclear transcription factor YY1. This factor regulates a multitude of cellular promoters, as an activator of transcription, as a repressor, or as an initiator element-binding protein. Antibodies directed against YY1 were used to show that YY1 copurified with the vaccinia late promoter-binding protein and was present in late promoter-protein complexes in gel supershift assays. Bacterially expressed YY1 also bound specifically to late promoter DNA. A dinucleotide replacement within the YY1 recognition motif directly adjacent to the transcription start site severely reduced the affinity of YY1 for the I1L promoter in vitro and impaired I1L promoter-dependent transcription in vivo. The intracellular localization of YY1 was shown by immunofluorescence microscopy to shift from primarily nuclear to the cytoplasm after vaccinia infection. These results indicate that YY1 has a positive role in the regulation of vaccinia virus late gene transcription and suggest that poxviruses have adapted cellular initiator elements as a means of regulating viral gene expression. This is the first identifiable cellular protein implicated in poxvirus transcription.

Base Sequence↗
PubMed1999

Extramedullary plasmacytoma: tumor occurrence and therapeutic concepts.

BACKGROUND: Extramedullary plasmacytoma (EMP) is a rare entity belonging to the category of non-Hodgkin lymphoma. EMPs make up 4% of all plasma cell tumors and occur mainly in the upper aerodigestive tract (UAD). Seven patients with EMP included in this evaluation were under the authors' care and have been clinically followed since 1990. Because there are no general guidelines for the treatment of patients with EMP, the authors tried to obtain detailed data about the occurrence of this disease and also reviewed the therapies that have been used. To do so, they evaluated all EMP cases published in the medical literature until now and included their own experience. METHODS: Based on the clinical course and follow-up of their own EMP patients, the authors evaluated and reinvestigated all EMP cases cited in MEDLINE, Index Medicus, DIMDI (Deutsches Institut fur medizinische Dokumentation und Information, Cologne, Germany), and the reference lists of the publications found through these sources. RESULTS: In a detailed literature search, more than 400 publications between 1905 and 1997 were found, and these revealed that EMP mainly occurs between the fourth and seventh decades of life. Seven hundred fourteen cases (82.2%) were found in the UAD, and 155 cases (17.8%) were found in other body regions. The following therapeutic strategies were used to treat patients with EMP of the UAD: radiation therapy alone in 44.3%, combined therapy (surgery and radiation) in 26.9%, and surgery alone in 21.9%. The median overall survival or recurrence free survival was longer than 300 months for patients who underwent combined intervention (surgery and radiation). This result was statistically highly significant (P = 0.0027, log rank test) compared with the results for patients who underwent surgical intervention alone (median survival time, 156 months) or radiation therapy alone (median survival time, 144 months). In most cases of non-UAD EMP, surgery was performed (surgery alone, 55.6%; surgery and radiation combined, 19.8%; radiation alone, 11.1%), but there were no statistical differences in survival (P = 0.62). Overall, after treatment for EMP in the UAD, 61.1% of all patients had no recurrence or conversion to systemic involvement (i.e., multiple myeloma, MM); however, 22.0% had recurrence of EMP, and 16.1% had conversion to MM. After treatment for EMP in non-UAD areas, 64.7% of all patients had no recurrence or MM, 21.2% had recurrence, and 14.1% had conversion to MM. CONCLUSIONS: The current investigation provides evidence that surgery alone gives the best results in cases of EMP of the UAD when resectability is good. However, if complete surgical tumor resection is doubtful or impossible and/or if lymph node areas are affected, then combined therapy (surgery and radiation) is recommended. These results, which were obtained from retrospective studies, should be confirmed in randomized trials comparing surgery with combined radiation therapy and surgery.

Adult↗
PubMed1999

Intratumoral heterogeneity in breast carcinoma revealed by laser-microdissection and comparative genomic hybridization.

To evaluate the potential cytogenetic heterogeneity in breast carcinoma, several small cell groups (each consisting of 20 to 50 cells) were investigated within paraffin sections. By laser-microdissection, three to seven cell groups were taken per case. The DNA was amplified by degenerate oligonucleotide primed PCR (DOP-PCR), and the samples were analyzed by CGH for chromosomal gains and losses. Two ductal invasive breast carcinomas, one of them with two lymphnode metastases, were investigated. To compare the results from the small samples, CGH was also performed on DNA isolated from the tumorous regions of three to five serial sections (10(7) to 10(6) cells). The aberrations observed in the microdissected tumor samples were multiple and involved up to 14 different chromosomal or subchromosomal regions. The most frequent changes were gains on chromosomes 12q (14/20) and 20q (16/20), and loss on 13q (12/20). Some aberrations have rarely been detected (e.g., loss on 2p, gain on 8q). Comparing chromosomal imbalances in primary tumors and lymph node metastases, more consistent changes were found between the primary tumor and its corresponding metastases than between both primary tumors. The laser-microdissected samples in general showed more chromosomal aberrations than DNA isolated from several tumor sections. Our CGH results were confirmed by fluorescence in situ hybridization (FISH) for the chromosomal regions of centromere 1 and 20, and 20q13. In addition, microsatellite analyses on 31 samples confirmed our CGH findings for selected chromosome regions 2p and 11q. It can be concluded that there is a distinct intratumoral heterogeneity in primary breast tumors as well as in the corresponding lymph node metastases. The combination of microdissection and CGH enabled us to detect cytogenetic aberrations from important clones which are missed when analyzing DNA extracted from large cell numbers.

Breast Neoplasms↗
PubMed1999

Androgens regulate bone resorption activity of isolated osteoclasts in vitro.

For many years it has been recognized that sex steroids have profound effects on bone metabolism. The current perception is that estrogen decreases bone resorption and androgen increases bone deposition. To investigate the potential for androgens to directly modulate bone resorption, we have examined avian osteoclast and human and mouse osteoclast-like cells for androgen responsiveness. There was a dose-dependent decrease in resorption activity in response to alpha-dihydrotestosterone (alpha-DHT), beta-DHT, testosterone, or the synthetic androgen RU1881. This decrease was blocked by cotreatment with the specific androgen antagonist hydroxyflutamide. Further examination of avian osteoclasts revealed that the cells exhibited specific and saturable nuclear binding of tritiated RU1881 and that alpha-DHT stimulated the activity of the androgen response element as measured by using a chloramphenicol acetyltransferase reporter plasmid. In addition, avian osteoclasts responded to androgen treatment with elevated production and secretion of transforming growth factor beta, a well documented response to androgen exposure in other cell systems. Treatment with either alpha-DHT or beta-DHT for 24 hours resulted in a significant dose-dependent decrease in secretion of cathepsin B and tartrate-resistant acid phosphatase. This response to beta-DHT, a stereoisomer of alpha-DHT that is inactive in other androgen receptor-dependent systems, supports the hypothesis that the osteoclast androgen receptor has unusual ligand-binding properties. Taken together, these results confirm the presence of functional androgen receptors in these cells and support the conclusion that osteoclasts are able to respond directly to androgens in vitro and thus are potential androgen target cells in vivo.

Acid Phosphatase↗
PubMed1999

[Techniques for postoperative monitoring of tissue circulation after free microvascular tissue transplantation].

Success rates after free tissue transplantation (FTT) have greatly improved over the last 20 years, partly due to improved technical performance of microvascular anastomoses with better optical and instrumental aids. However, flap failure is still a clinical problem and occurs in 5 to 10%, mainly due to blood vessel thrombosis within the first 24 postoperative hours. The clinical results after FTT can be optimized by in-time diagnosis of irreversibly compromised tissue blood flow and immediate operative reexploration. Therefore, there is a special demand for adequate and reliable postoperative monitoring techniques. This article reviews all monitoring techniques which have been performed in the experimental-clinical setting after FTT thus far.

Follow-Up Studies↗
PubMed1998

Changes in the properties of gap junctions during neuronal differentiation of hippocampal progenitor cells.

The cellular mechanisms that regulate progenitor cell lineage elaboration and maturation during embryonic development of the mammalian brain are poorly understood. Conditionally immortalized mouse hippocampal multipotent progenitor cells (MK31 cells) were found to be strongly coupled by gap junctions comprising connexin 43 (Cx43) during early neuronal ontogeny; the presence of this Cx type was confirmed by electrophysiological, molecular biological, and immunocytochemical assays. However, as progenitor cells underwent intermediate stages of neuronal differentiation under the influence of interleukin 7 (IL-7) alone or terminal differentiation after composite exposure to basic fibroblast growth factor, IL-7, and transforming growth factor alpha, coupling strength and the level of Cx43 expression declined. An additional population of junctional channels with distinct properties was detected at an intermediate stage of neuronal differentiation. Reverse transcription-PCR assays detected mRNA encoding Cx40 in IL-7-treated cells and Cx33 after both treatment conditions. Because functional channels in exogenous expression systems are not formed by pairing Cx40 with Cx43 or by pairing Cx33 with itself or additional connexins, these experimental observations raise the possibility that the progressive loss of coupling during differentiation of neural progenitor cells may involve downregulation of Cx43 coupled with potentiation of expression of Cx33 and Cx40. Furthermore, continued expression of Cx43 in differentiating neuroblasts could mediate intercellular communication between neuronal precursor cells and astrocytes by direct signaling via homotypic gap junction channels.

Animals↗
PubMed1998

The effect of changing sexual activity on HIV prevalence.

In a one-sex preferred mixing model, reductions in the rate of partner change by those with low sexual activity increase the average probability of HIV infection in the remaining pool of available partners. This increases prevalence among people with high activity, and since high activity people disproportionately influence the spread of HIV, may increase long-run prevalence in the population as a whole. Calculations using the model and survey data on sexual activity indicate that in low prevalence populations, many people have low enough activity that reductions in their activity might increase the endemic steady-state prevalance. If these results prove robust in more realistic models, they would support the case for targeting public health messages urging reduced sexual activity to high activity people.

Computer Simulation↗
PubMed1998

Dye coupling between spinal cord oligodendrocytes: differences in coupling efficiency between gray and white matter.

Oligodendrocytes express two gap junction proteins, connexin32 (Cx32) and Cx45. To test for functional coupling between oligodendrocytes, cells were filled with the (Cx32-permeable) dyes Lucifer Yellow (LY) and Neurobiotin. Cells in slices from rat spinal cord were dialyzed via the patch pipette containing the dye while recording with the patch-clamp technique. The dye-labeled cells were identified as oligodendrocytes by their characteristic pattern of membrane currents and by morphology. In gray matter, 18% of the injected cells (N = 94) were coupled to more than three adjacent cells (slices from postnatal day 1 to 19). In contrast, in white matter, the dye was restricted to the injected cell (N = 63 for Lucifer Yellow injection only; N = 11 for LY and Neurobiotin) indicating a lack of functional coupling. Immunolabeling of Cx32 in mature oligodendrocytes of white matter revealed that the gap junction protein is localized on the cell bodies and abaxonal processes which occupy non-overlapping territories. In immature white matter and gray matter, Cx32 is mostly concentrated in the somatic region of the cells. In addition to Cx32, we have obtained immunocytochemical data that oligodendrocytes can express Cx45 with a labeling pattern different from the Cx32 expression. Two alternative interpretations of the coupling data are discussed: 1) that the presence of Cx32 in mature white matter oligodendrocytes does not serve for communication between cells, but rather for communication within oligodendrocytes in the sense of autocellular coupling, or 2) that the glial syncytium is furnished with a high degree of functional rectification at the oligodendrocytic side.

Animals↗
PubMed1998

The National Practitioner Data Bank: implications for nurse anesthetists.

The National Practitioner Data Bank (NPDB), created by the 1986 Health Care Quality Improvement Act, has been in operation since 1990. Hospitals and other credentialing bodies must query the NPDB when granting and renewing privileges. The NPDB receives about 25,000 reports of adverse actions against health practitioners each year. The NPDB was designed to be a flagging system providing information to licensing or credentialing authorities who would further examine practitioner records. Its purpose is to ensure that decision makers have information that might not otherwise be readily available, especially in the case of incompetent practitioners who move from hospital to hospital or state to state. Access to NPDB information is a concern for consumers and providers alike. Only 2% of matched reports to the NPDB made a difference in hospital privileging decisions. A limitation of NPDB information is that malpractice payments recorded in the NPDB do not necessarily constitute a comprehensive and definitive reflection of actual health care incompetence. All health care providers need to be aware of the NPDB, its mission, potential impact on their ability to be credentialed, and proposed additional uses of its information.

Clinical Competence↗
PubMed1997

Expression of connexin43 mRNA in adult rodent brain.

The expression of connexin43 mRNA was detected in adult rat brains by in situ hybridization methods. Specific digoxigenin riboprobes were generated by in vitro transcription of two PCR-amplified fragments of connexin43 cDNA which lack homology with any other published connexin. Following immunohistochemical detection, the digoxigenin cRNA was found to occur in various neuronal populations including Purkinje cells of the cerebellum, pyramidal cells of the neocortex and the hippocampal formation, as well as granule cells of the dentate gyrus and various neurons of diverse hindbrain nuclei. This ubiquituous expression of connexin43 mRNA in adult neurons, in particular in neocortical pyramidal cells, is surprising insofar as gap junction communication in adult bains has been considered to be confined to specific subpopulations of neurons revealing a high incidence of synchronized electrical activities in contrast to the postnatal brain where interneuronal coupling via gap junctions precedes the formation of chemical transmission. In addition, connexin43 is regarded as being preferentially expressed in astrocytes, although its presence in adult neurons has not definitely been excluded. We propose that adult neurons preserve their capability of expressing functional gap junctions more frequently than presently considered and that connexin43 is a most likely neuronal gap junction protein candidate.

Animals↗