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Biomedical subjects

M Pfeffer

Publications and source records attributed to M Pfeffer.

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

Antisense experiments reveal molecular details on mechanisms of ICER suppressing cAMP-inducible genes in rat pinealocytes.

In the rat pineal gland neuronal signals determine the rhythmic synthesis of the hormone melatonin. Norepinephrine (NE) is the principal neurotransmitter that drives hormone synthesis by activating the cAMP signaling pathway. This activation depends on transcriptional and posttranscriptional regulatory mechanisms. The cAMP-dependent transcriptional regulation of the rate-limiting enzyme of melatonin synthesis, arylalkylamine-N-acetyltransferase (AA-NAT) involves the activating transcription factor (TF) CREB and the inhibitory TF ICER. By silencing elements of this cAMP-dependent neuroendocrine transduction cascade we wished to gain further insight into the role of ICER in the regulation of gene expression in rat pineal gland. Inhibition of specific kinases in primary pinealocyte cultures showed that ICER induction depends pivotally on the activation of cAMP-dependent protein kinase II. Eliminating ICER's impact by transfecting antisense constructs into pinealocytes revealed a predominant beta-adrenergic mechanism in regulating a cotransfected CRE-inducible reporter gene and notably, also the endogenous AA-NAT gene. Deciphering molecular details of the cAMP-dependent gene expression in mammalian pinealocytes provides a basis for understanding the general architecture of this signaling pathway that serves adaptive processes ubiquitously in the organism.

Animals↗
PubMed2000

Development of reverse transcription-PCR assays specific for detection of equine encephalitis viruses.

Specific and sensitive reverse transcription-PCR (RT-PCR) assays were developed for the detection of eastern, western, and Venezuelan equine encephalitis viruses (EEE, WEE, and VEE, respectively). Tests for specificity included all known alphavirus species. The EEE-specific RT-PCR amplified a 464-bp region of the E2 gene exclusively from 10 different EEE strains from South and North America with a sensitivity of about 3,000 RNA molecules. In a subsequent nested PCR, the specificity was confirmed by the amplification of a 262-bp fragment, increasing the sensitivity of this assay to approximately 30 RNA molecules. The RT-PCR for WEE amplified a fragment of 354 bp from as few as 2,000 RNA molecules. Babanki virus, as well as Mucambo and Pixuna viruses (VEE subtypes IIIA and IV), were also amplified. However, the latter viruses showed slightly smaller fragments of about 290 and 310 bp, respectively. A subsequent seminested PCR amplified a 195-bp fragment only from the 10 tested strains of WEE from North and South America, rendering this assay virus specific and increasing its sensitivity to approximately 20 RNA molecules. Because the 12 VEE subtypes showed too much divergence in their 26S RNA nucleotide sequences to detect all of them by the use of nondegenerate primers, this assay was confined to the medically important and closely related VEE subtypes IAB, IC, ID, IE, and II. The RT-PCR-seminested PCR combination specifically amplified 342- and 194-bp fragments of the region covering the 6K gene in VEE. The sensitivity was 20 RNA molecules for subtype IAB virus and 70 RNA molecules for subtype IE virus. In addition to the subtypes mentioned above, three of the enzootic VEE (subtypes IIIB, IIIC, and IV) showed the specific amplicon in the seminested PCR. The practicability of the latter assay was tested with human sera gathered as part of the febrile illness surveillance in the Amazon River Basin of Peru near the city of Iquitos. All of the nine tested VEE-positive sera showed the expected 194-bp amplicon of the VEE-specific RT-PCR-seminested PCR.

Alphavirus↗
PubMed2000

Cellular localization of fibroblast growth factor 2 (FGF-2) in benign prostatic hyperplasia.

Fibroblast growth factor 2 (FGF-2, basic fibroblast growth factor) has been reported to be elevated in tissues from benign prostatic hyperplasia (BPH), the most frequent neoplastic disease in aging men. This suggests that FGF-2 may play a significant role in the development of BPH. In this study the cellular distribution pattern of FGF-2 in tissues from BPH has been investigated by immunohistochemical and molecular biological methods. Radioimmunoassay revealed high concentrations of FGF-2, ranging between 450 and 950 ng per g tissue. Immunoblots confirmed the presence of a 18 kDa FGF-2 in tissue extracts. By immunohistochemistry done with a polyclonal antibody to recombinant FGF-2 on paraffin sections, FGF-2 was localized in fibroblasts, endothelial cells and smooth muscle cells of tissue samples of BPH. Nuclei of these cells were labelled distinctly. Moreover the cytoplasm of smooth muscle cells was labelled moderately. No immunostaining was seen in prostatic epithelium. Non-radioactive in situ hybridization with digoxygenin-labelled oligonucleotides revealed the presence of mRNA for FGF-2 in smooth muscle cells of the prostatic stroma. These results provide evidence that FGF-2 may be produced locally in the human prostate as a stroma-specific mitogen and may play a causal role in the development of BPH.

Fibroblast Growth Factor 2↗
PubMed2000

[Other side of the problem: "Systemic solutions of sexual functional disturbances"].

From a systemic constructive point of view it seems to be obvious that symptoms of any kind are to be looked at as definite human communicational behaviour. From this angle the sexual behaviour between two or more autonomous (self-conscious) humans can be defined as dialogue. Therefore, sexual problems can be understood, analysed and dealt with as communication problems. A very important and therapeutic idea is to consider symptoms as intervention for relationships and possibility for problem solutions. Furthermore, these disturbances are not to be considered as deficiency but as efficiency and competence. The consequences of this therapeutic attitude in a setting are explained in theory and practice as follows.

Couples Therapy↗
PubMed2000

Impairment in quality of life among patients seeking surgery for hyperhidrosis (excessive sweating): preliminary results.

BACKGROUND: The present paper describes the initial stages of the development and administration of a short, disease-specific, health related questionnaire to assess the impact of suffering from hyperhidrosis (excessive sweating) on the Quality of Life (QoL) of patients who are anticipating surgery for this disorder. METHOD: The study was performed in two stages: 1. The life domains in which the condition impairs QoL were assessed by in-depth interviews with 10 patients suffering from hyperhidrosis. 2. A questionnaire covering five life domains was built based on these interviews. 3. This questionnaire was administered to 48 patients, 30 females and 18 males between the ages 15 and 48. RESULTS: Results showed that subjective QoL was significantly lower among females in four of the five life areas and that duration of the condition correlates with a lower quality of life. A regression analysis showed that the subjective suffering of the patients was explained mainly by social aspects. CONCLUSIONS: The questionnaire is a novel attempt to assess QoL in a disorder with strong esthetic and social consequences and could improve communication between patients and their physicians.

Adult↗
PubMed2000

Mousepox resulting from use of ectromelia virus-contaminated, imported mouse serum.

Mousepox was identified in a single mouse-holding room in early 1999 after a group of 20 CAF1/Hsd mice were inoculated SC with a killed murine spindle cell tumor line, S1509A. The cell line had been used without complications multiple times and was determined to be free of viral contamination on the basis of results of mouse antibody production testing. Of the 20 mice inoculated, 12 mice died by postinoculation day 8. Severe lymphoid and hepatic necrosis was observed in select mice subjected to histologic examination. Ballooning degeneration of epithelial cells with intracytoplasmic eosinophilic inclusion bodies was observed in the skin overlying the inoculation site of the single mouse from which this tissue site was evaluated. Presence of ectromelia virus was confirmed by use of immunohistochemical and polymerase chain reaction analyses, and the virus was isolated after serum, pooled from 5 of the index cases, was inoculated into an immune-naive mouse. Investigation into the source of virus contamination included inoculating mice with aliquots of various S1509A freeze dates; chemically defined media and supplements, including fetal bovine serum; and two lots of pooled commercial mouse sera, after heat inactivation at 56 degrees C for 30 minutes used as a medium supplement. One lot of pooled commercial mouse serum was identified as the source of ectromelia virus. This lot of serum was inadvertently used to feed S1509A cells that were subsequently inoculated into mice. We determined that the contaminated serum, which was purchased in late 1998, originated from China. The serum was imported into the United States as a batch of 43 L in early 1995. The serum was blended into a single lot and filtered (0.2 microm) before distribution to major suppliers throughout the country. The serum was sold or further processed to obtain a variety of serum-derived products. Because murine serum is generally sold in small aliquots (10 to 50 ml), we speculate that several thousand aliquots may have been derived from this batch of serum and, if inoculated into mice, would likely result in additional mousepox outbreaks.

Animals↗
PubMed1999

[Persistent arthralgias in Ross-River-Virus disease after travel to the South Pacific].

HISTORY AND CLINICAL FINDINGS: A 57-year-old patient presented with malaise and severe persistent arthralgia of the left shoulder. He reported an acute illness with fever, generalized myalgia and arthralgias of the large joints which had started one month earlier during his flight back to Germany after a two weeks trip to the South Pacific. Physical examination showed extensive pain on palpation of the glenohumeral and acromioclavicular joints with impairment of active and passive mobility. Investigation of the cervical spine was normal. INVESTIGATIONS: Apart from elevated C-reactive protein and erythrocyte sedimentation rate levels, routine laboratory investigations were normal including negative immunodiagnostic tests for autoantibodies and various global infections that may be associated with arthritis. Immunofluorescence tests showed significant levels of specific IgM- and IgG-antibodies against Ross River virus (RRV) but not against other arboviruses endemic in the South Pacific and Australia (Dengue, West Nile, Chikungunya, Sindbis, Barmah Forest). This was confirmed by a positive RRV neutralisation test. Attempts at virus isolation and detection of viral RNA by PCR were not successful. TREATMENT AND COURSE: Symptomatic treatment with high doses of diclofenac quickly led to pain relief, and arthralgias receded within 10 days after begin of treatment. However, several bouts of arthralgia of the left shoulder and left knee occurred during a period of 4 months. CONCLUSIONS: Because of the current epidemiological situation in the South Pacific and Australia, infections by arboviruses like RRV should be considered in travellers returning from these areas with severe and persistent arthralgia of unknown origin, even in the absence of fever and other symptoms of acute infection.

Alphavirus Infections↗
PubMed1999

Transcription factors in neuroendocrine regulation: rhythmic changes in pCREB and ICER levels frame melatonin synthesis.

Neurotransmitter-driven activation of transcription factors is important for control of neuronal and neuroendocrine functions. We show with an in vivo approach that the norepinephrine cAMP-dependent rhythmic hormone production in rat pineal gland is accompanied by a temporally regulated switch in the ratio of a transcriptional activator, phosphorylated cAMP-responsive element-binding protein (pCREB), and a transcriptional inhibitor, inducible cAMP early repressor (ICER). pCREB accumulates endogenously at the beginning of the dark period and declines during the second half of the night. Concomitant with this decline, the amount of ICER rises. The changing ratio between pCREB and ICER shapes the in vivo dynamics in mRNA and, thus, protein levels of arylalkylamine-N-acetyltransferase, the rate-limiting enzyme of melatonin synthesis. Consequently, a silenced ICER expression in pinealocytes leads to a disinhibited arylalkylamine-N-acetyltransferase transcription and a primarily enhanced melatonin synthesis.

Animals↗
PubMed1999

Signal transduction in the rodent pineal organ. From the membrane to the nucleus.

The rodent pineal organ transduces a photoneural input into a hormonal output. This photoneuroendocrine transduction leads to highly elevated levels of the hormone melatonin at night-time which serves as a message for darkness. The melatonin rhythm depends on transcriptional, translational and posttranslational regulation of the arylalkylamine-N-acetyltransferase, the key enzyme of melatonin biosynthesis. These regulatory mechanisms are fundamentally linked to two second messenger systems, namely the cAMP- and the Ca(2+)-signal transduction pathways. Our data gained by molecular biology, immunohistochemistry and single-cell imaging demonstrate a time- and substance-specific activation of these signaling pathways and provide a framework for the understanding of the complex signal transduction cascades in the rodent pineal gland which in concert not only regulate the basic profile but also fine-tune the circadian rhythm in melatonin synthesis.

Animals↗
PubMed1999

Characterization of orthopoxviruses isolated from man and animals in Germany.

Fourteen orthopoxvirus strains isolated from humans, cats, a dog, a cow, and an elephant in Germany were characterized. All were classified as cowpox virus based on haemorrhagic lesions induced on the chorioallantoic membrane of chicken eggs and reactivity of a 160 kDa protein with anti-A-type inclusion protein hyperimmune serum in a Western blot. More detailed comparison of the isolates by restriction endonuclease mapping using HindIII and XhoI demonstrated a close relationship between all isolates and confirmed them as cowpox viruses. However, some minor differences between the isolates were detected which proved to be of epidemiological value. One group consisting of five closely related isolates contained a unique 4.0 kb HindIII fragment. In a Southern blot this fragment failed to hybridize with other cowpox virus isolates including the reference strain.

Amino Acid Sequence↗
PubMed1999

Sequencing of prototype viruses in the Venezuelan equine encephalitis antigenic complex.

The 5' nontranslated region (5'NTR) and nonstructural region nucleotide sequences of nine enzootic Venezuelan equine encephalitis (VEE) virus strains were determined, thus completing the genomic RNA sequences of all prototype strains. The full-length genomes, representing VEE virus antigenic subtypes I-VI, range in size from 11.3 to 11.5 kilobases, with 48-53% overall G+C contents. Size disparities result from subtype-related differences in the number and length of direct repeats in the C-terminal nonstructural protein 3 (nsP3) domain coding sequence and the 3'NTR, while G+C content disparities are attributable to strain-specific variations in base composition at the wobble position of the polyprotein codons. Highly-conserved protein components and one nonconserved protein domain constitute the VEE virus replicase polyproteins. Approximately 80% of deduced nsP1 and nsP4 amino acid residues are invariant, compared to less than 20% of C-terminal nsP3 domain residues. In two enzootic strains, C-terminal nsP3 domain sequences degenerate into little more than repetitive serine-rich blocks. Nonstructural region sequence information drawn from a cross-section of VEE virus subtypes clarifies features of alphavirus conserved sequence elements and proteinase recognition signals. As well, whole-genome comparative analysis supports the reclassification of VEE subtype-variety IF and subtype II viruses.

5' Untranslated Regions↗
PubMed1999

Candesartan in heart failure--assessment of reduction in mortality and morbidity (CHARM): rationale and design. Charm-Programme Investigators.

BACKGROUND: Chronic heart failure (CHF) is an increasing burden to health care. Pharmacological treatment with angiotensin-converting enzyme (ACE) inhibitors and beta blockers improve survival and reduce hospitalizations in patients with low left ventricular ejection fraction (LVEF). Despite these therapies, morbidity and mortality remains problematic. Furthermore, 30% to 50% of patients with CHF have a preserved LVEF. It is not known if treatments are of benefit in this group. DESIGN: Candesartan in Heart Failure-Assessment of Reduction in Mortality and Morbidity (CHARM) is a program designed to investigate the clinical usefulness of the long-acting angiotensin II type 1 receptor blocker, candesartan cilexetil, in a broad spectrum of patients with symptomatic heart failure. Patients with systolic dysfunction, tolerant or intolerant to an ACE-inhibitor, and patients with preserved systolic function are included. Specifically, the CHARM program consists of 3 independent, parallel, placebo-controlled studies in patients with (1) LVEF less than or equal to 40%, ACE-inhibitor treated (n = 2,300); (2) LVEF less than or equal to 40%, ACE-inhibitor intolerant (n = 1,700); (3) LVEF greater than 40%, not treated with ACE inhibitors (n = 2,500). The 3 studies will be combined to evaluate the effect of candesartan cilexetil on all-cause mortality in the broad spectrum of symptomatic heart failure. The primary objective in each trial is to evaluate the effects on the combined endpoint of cardiovascular mortality or CHF hospitalization. Other endpoints include the effects on myocardial infarction, all-cause hospitalization, and resource utilization. CHARM is intended to randomize 6,500 patients with symptomatic heart failure from 26 countries in Europe, the United States, Canada, South Africa, and Australia. The CHARM program started to enroll patients in March 1999. The follow-up period is a minimum of 2 years. The study is expected to end in the third quarter of 2002.

Adolescent↗
PubMed1999

Inducible cyclic AMP early repressor protein in rat pinealocytes: a highly sensitive natural reporter for regulated gene transcription.

Rhythmic activity of arylalkylamine N-acetyltransferase (AANAT) determines melatonin synthesis in rat pineal gland. The transcriptional regulation of AANAT involves the activating and inhibiting transcription factors of the cyclic AMP (cAMP)-signaling pathway, cAMP response element-binding protein and inducible cAMP early repressor (ICER), respectively. Activation of this pathway is centered around norepinephrine, stimulating beta(1)-adrenergic receptors, but various other transmitters can modulate melatonin biosynthesis. To compare the transcriptional impact of norepinephrine with that of other neurotransmitters on melatonin synthesis, we determined ICER protein levels in pinealocytes and, in parallel, hormone secretion. The dose-dependent inductions of ICER protein by norepinephrine, the beta(1)-adrenergic receptor agonist isoproterenol, vasoactive intestinal peptide, pituitary adenylate cyclase-activating polypeptide, and adenosine are correlated to regulatory dynamics in melatonin production. Importantly, ICER protein induction required lower ligand concentrations than the induction of melatonin biosynthesis. Although neuropeptide Y, glutamate, and vasopressin altered norepinephrine-stimulated hormone production without affecting ICER levels, the activation of voltage-gated cation channels increased ICER without affecting hormone synthesis. Sensitivity and versatility of ICER induction in pinealocytes make these neuroendocrine cells a valuable model system in which to study molecular interactions determining a regulated gene expression.

Acetylcholine↗
PubMed1999

Genetic evidence for the origins of Venezuelan equine encephalitis virus subtype IAB outbreaks.

Epizootics of Venezuelan equine encephalitis (VEE) involving subtype IAB viruses occurred sporadically in South, Central and North America from 1938 to 1973. Incompletely inactivated vaccines have long been suspected as a source of the later epizootics. We tested this hypothesis by sequencing the PE2 glycoprotein precursor (1,677 nucleotides) or 26S/nonstructural protein 4 (nsP4) genome regions (4,490 nucleotides) for isolates representing most major outbreaks. Two distinct IAB genotypes were identified: 1) 1940s Peruvian strains and 2) 1938-1973 isolates from South, Central, and North America. Nucleotide sequences of these two genotypes differed by 1.1%, while the latter group showed only 0.6% sequence diversity. Early VEE virus IAB strains that were used for inactivated vaccine preparation had sequences identical to those predicted by phylogenetic analyses to be ancestors of the 1960s-1970s outbreaks. These data support the hypothesis of a vaccine origin for many VEE outbreaks. However, continuous, cryptic circulation of IAB viruses cannot be ruled out as a source of epizootic emergence.

Amino Acid Sequence↗
PubMed1999

[Cowpox viruses in Germany: an analysis of 5 cases in 1998].

Five case reports on cowpox virus infections in cats, humans, and for the first time in a horse are presented. It becomes obvious that in most cases the diagnosis cowpox is suspected rather late, although fast and reliable diagnostic tools such as pathohistological examination and polymerase chain reaction are available. The threat of a zoonotic transmission mainly through cats is gaining importance. Although wild rodents have been claimed to be the reservoir and source for cowpox viruses in cats, very little is known about the epidemiology of cowpox virus. Based on the different genome organizations found in the German isolates, we conclude that various subtypes of cowpox virus are circulating in Germany at one time.

Animals↗
PubMed1998

Ontogeny of a diurnal rhythm in arylalkylamine-N-acetyltransferase mRNA in rat pineal gland.

Melatonin synthesis in the pineal gland of adult rats is linked to cAMP-dependent transcriptional and post-transcriptional regulatory mechanisms affecting its rate-limiting enzyme, the arylalkylamine-N-acetyltransferase (AA-NAT). During development of the pineal gland, neuronal control gains access to the earlier matured cAMP-signaling pathway to shape the day-night rhythm in AA-NAT enzymatic activity. By semiquantitative in situ hybridization we analyzed if the developmental onset of a rhythmic AA-NAT activity is correlated to a temporally parallel onset in AA-NAT transcription. We found that AA-NAT mRNA levels in rat pineal gland become rhythmic at postnatal day 5. Thus, AA-NAT gene transcription in rat pineal gland starts to show day-night differences shortly prior to the appearance of a rhythmic AA-NAT activity.

Animals↗
PubMed1998

The alphavirus 3'-nontranslated region: size heterogeneity and arrangement of repeated sequence elements.

The 3'-nontranslated region (NTR) of representative strains of all known alphavirus species was amplified by reverse transcription-polymerase chain reaction. For 23 of them, the 3'-NTR sequence was determined. Together with previously published data, this allowed an analysis of the 3'-NTR of the viruses in the genus Alphavirus. The length of the 3'-NTRs varied from 77 nt for Pixuna virus to 609 nt for Bebaru virus. The 19-nt conserved sequence element directly adjacent to the poly(A) tract was found in all viruses, supporting the hypothesis that this region is a cis-acting sequence element during viral replication and essential for virus growth in vitro. Within the 3'-NTR of all alphaviruses, repeated sequence elements of various numbers and lengths were found. Their composition was very consistent in both the Venezuelan equine encephalitis (VEE) and the Sindbis-like viruses, although their number was constant only within the latter group. For the VEE viruses, our data suggested that insertion events rather than deletions from an ancestor with a long 3'-NTR created the various number of repeated sequence elements. Among the remaining viruses, both the number and the composition of repeated sequence elements varied remarkedly.

Alphavirus↗