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Biomedical subjects

K Frank

Publications and source records attributed to K Frank.

At least 55 records · Page 3Linked to original sources
PubMed1997

Expression of tenascin-C by human endometrial adenocarcinoma and stroma cells: heterogeneity of splice variants and induction by TGF-beta.

Localization of tenascin-C in vivo and cell culture experiments in vitro have provided evidence for stromal production of tenascin-C in malignant tumors of a variety of organs. Here we raised the question of whether the mesenchymal stroma in the case of endometrial adenocarcinoma is the unique source of tenascin-C. Therefore, the expression of tenascin-C mRNA by human endometrial adenocarcinoma cells and endometrial stroma cells was investigated. Several preparations of endometrial stroma cells produced tenascin-C mRNA. Using a serum-free defined cell culture medium, production of tenascin-C mRNA could be increased by adding either serum or 20 ng TGF-beta/mL to the cell culture medium. Reverse transcriptase polymerase chain reaction analysis revealed that five out of six endometrial adenocarcinoma cell lines produced tenascin-C mRNA. Northern blot experiments and ribonuclease protection assays provided evidence that the number of copies of tenascin-C mRNA was small. Analysis of expressed splice variants by reverse transcriptase polymerase chain reaction analysis revealed the abundance of one major splice variant that lacked all potential alternatively spliced fibronectin type-III-like repeats. Regarding larger splice variants, all fragment sizes that could theoretically originate from seven alternatively spliced fibronectin type-III-like repeats were observed. Evaluating relative signal intensities, the splice variants containing a single fibronectin type-III-like repeat and the variant possessing all but one alternatively spliced repeats were most frequent. In summary, evidence is provided that tenascin-C can originate from both tissue compartments of the human endometrium stroma and (tumor) epithelium. Splice variant analysis revealed a high number of splice variants and a relative high proportion of variants that have so far been regarded as minor constituents of expressed tenascin-C.

Adenocarcinoma↗
PubMed1996

Induction of the membrane alanyl aminopeptidase gene and surface expression in human T-cells by mitogenic activation.

The metal-dependent membrane alanyl aminopeptidase (amino-peptidase N, APN, CD13; EC 3.4.11.2) is a well-established marker of normal and malignant cells of the myelo-monocytic lineage. It is also expressed by leukaemic blasts of a small group of patients suffering from acute or chronic lymphoid leukaemia. CD13-specific monoclonal antibodies do not bind to the surface of normal B lymphocytes, and APN mRNA was not detectable by Northern analysis in normal lymphocytes or in T-cell lines. Recently the expression of the APN gene in T-cell lines as well as the ability of these cells to cleave chromogenic substrates preferred by APN have been demonstrated [Lendeckel, Wex, Kähne, Frank, Reinhold and Ansorge (1994) Cell. Immunol. 153, 214-226]. Here, by means of dot-blot hybridization and RNase protection assay, evidence is provided that human peripheral T-cells as well as derived cell lines contain significant amounts of APN mRNA, comparable to that in the promyeloic cell line U937, and that mitogenic activation of peripheral human T-cells leads to a more than 4-fold increase in their APN mRNA content. In the course of activation, T-cells increase their total alanine p-nitroanilide-hydrolysing activity to approx. 7-fold that of resting cells. Furthermore these cells become immunoreactive towards CD13 to a significant extent (up to 51%) as shown by surface staining and confirmed by activity staining and immunostaining after isoelectric focusing (pI of T-cell APN = 4.6). In addition it is demonstrated by fluorescence microscopy that viable, activated T-cells effectively cleave the fluorogenic aminopeptidase substrate bis-glycyl-rhodamine 110 and that the corresponding aminopeptidase activity is associated with the cell surface. We show that specific inhibitors of APN, probestin and actinonin, strongly decrease DNA synthesis in phytohaemagglutinin (PHA)-stimulated T-cells. In summary, evidence is presented that in the course of mitogenic activation human peripheral T-cells increase the expression of APN both at the transcriptional level and at the cell surface. This has been demonstrated both at the APN mRNA level and at the protein level with respect to aminopeptidase enzymic activity and CD13 immunoreactivity.

CD13 Antigens↗
PubMed1996

Regional expression of sodium pump subunits isoforms and Na+-Ca++ exchanger in the human heart.

Cardiac glycosides exert a positive inotropic effect by inhibiting sodium pump (Na,K-ATPase) activity, decreasing the driving force for Na+-Ca++ exchange, and increasing cellular content and release of Ca++ during depolarization. Since the inotropic response will be a function of the level of expression of sodium pumps, which are alpha(beta) heterodimers, and of Na+-Ca++ exchangers, this study aimed to determine the regional pattern of expression of these transporters in the heart. Immunoblot assays of homogenate from atria, ventricles, and septa of 14 nonfailing human hearts established expression of Na,K-ATPase alpha1, alpha2, alpha3, beta1, and Na+-Ca++ exchangers in all regions. Na,K-ATPase beta2 expression is negligible, indicating that the human cardiac glycoside receptors are alpha1beta1, alpha2beta1, and alpha3beta1. alpha3, beta1, sodium pump activity, and Na+-Ca++ exchanger levels were 30-50% lower in atria compared to ventricles and/or septum; differences between ventricles and septum were insignificant. Functionally, the EC50 of the sodium channel activator BDF 9148 to increase force of contraction was lower in atria than ventricle muscle strips (0.36 vs. 1.54 microM). These results define the distribution of the cardiac glycoside receptor isoforms in the human heart and they demonstrate that atria have fewer sodium pumps, fewer Na+-Ca++ exchangers, and enhanced sensitivity to inotropic stimulation compared to ventricles.

Adult↗
PubMed1996

Evidence for functional relevance of an enhanced expression of the Na(+)-Ca2+ exchanger in failing human myocardium.

BACKGROUND: The present study aimed at investigating the expression of the Na(+)-Ca2+ exchanger and its functional role in human failing myocardium. METHODS AND RESULTS: Na(+)-Ca2+ exchanger mRNA and protein levels were examined in nonfailing (NF, n = 8) and failing human myocardium (New York Heart Association functional class IV) with idiopathic dilated cardiomyopathy (DCM, n = 8) or ischemic heart disease (ICM, n = 6). The inotropic effect of the Na+ channel activator BDF 9148 was determined in electrically driven left ventricular papillary muscle strip preparations (1 Hz, 37 degrees C) from nonfailing (n = 8) and failing (n = 8) human hearts. Na(+)-Ca2+ exchanger mRNA levels were significantly increased, by 79% (P < .001) in DCM and by 58% (P < .01) in ICM compared with NF; protein levels increased by 36% (P < .001) and by 20% (P < .05), respectively. BDF 9148 increased the force of contraction concentration dependently, with a similar maximal effect in NYHA class IV and NF, but was more potent in NYHA class IV as demonstrated by a significantly smaller (P < .01) EC50 value (NYHA class IV, 0.18 [0.16 to 0.22] mumol/L; NF, 1.65 [1.3 to 3.0] mumol/L). In NYHA class IV, BDF 9148 (0.1 mumol/L) restored the positive force-frequency relationship and reduced the frequency-dependent increase in diastolic tension in relation to force of contraction. CONCLUSIONS: The increased expression of the Na(+)-Ca2+ exchanger is a possible explanation for the increased inotropic potency of the Na+ channel activator BDF 9148 in failing human myocardium. The increase in exchanger molecules could be of functional relevance for the modulation of cardiac contractility by agents that increase the intracellular Na+ concentration. Enhancement of Na(+)-Ca2+ exchanger activity might be a powerful mechanism for increasing cardiac contractility in chronic heart failure.

Adult↗
PubMed1996

Altered inotropism in the failing human myocardium.

Beta-adrenoreceptor-cAMP-dependent inotropic interventions lose their effectiveness depending on the degree of myocardial failure. This blunted effect of beta-adrenoreceptor-dependent stimulation might be due to a downregulation of beta-adrenoreceptors and an increase of inhibitory G-proteins leading to decreased intracellular cAMP-concentrations. However, the maximal positive inotropic effect elicited by elevation of the extracellular [Ca2+] does not differ between failing and nonfailing human myocardium, indicating that terminally failing human myocardium is effective to increase force of contraction to the same degree as nonfailing tissue. Agents which increase force of contraction primarily via increasing the intracellular [Na+], e.g., cardiac glycosides and the Na(+)-channel activator BDF 9148, exert a higher potency in failing myocardium than in nonfailing tissue to increase force of contraction. This could result from an enhanced protein expression of the Na+/Ca(2+)-exchanger observed in diseased human hearts. Alterations in the intracellular Ca(2+)-homeostasis reported in failing myocardium lead to a negative force-frequency-relationship and a prolonged relaxation. As the protein expression of SERCA IIa and phospholamban seems to be similar in NYHAIV and nonfailing tissue, the reduced Ca(2+)- uptake may result from an altered regulation of these proteins, e.g., reduced phosphorylation of phospholamban or the SERCA IIa. After inhibition of the Ca(2+)-ATPase of the sarcoplasmic reticulum with the high specific inhibitor cyclopiazonic acid the former positive force-frequency-relationship became significantly less positive even in the nonfailing tissue and twitch course became similar to diseased hearts. These findings may be indicative for the importance of the Ca(2+)-reuptake mechanism into the sarcoplasmic reticulum in addition to the regulatory control at the site of the contractile apparatus for the regulation of contraction and relaxation in human myocardium.

Adrenergic beta-Agonists↗
PubMed1996

Enhanced sensitivity of the failing human myocardium to cardiac glycosides and Na(+)-channel activators.

Cardiac glycosides and Na+ -channel activators increase intracellular Na+ and thereby enhance the transport rate of the sarcolemmal Na+/Ca2+ exchanger. We tested the hypothesis of whether increased expression of the Na+/Ca2+ exchanger in failing human myocardium is accompanied by enhanced sensitivity of the failing human myocardium toward cardiac glycosides and Na+ -channel activators. We studied the positive inotropic effects of the new Na+ -channel activator BDF and the cardiac glycoside ouabain in human failing (New York Heart Association [NYHA] functional class IV, heart transplants for dilated cardiomyopathy, n = 11) and nonfailing (donor hearts, n = 5) myocardium on electrically driven left ventricular papillary muscle strips (1 Hz, 37 degrees C). The effectiveness of ouabain and BDF to increase force of contraction was similar in human nonfailing and failing myocardium. BDF was more potent to increase force of contraction in failing than in nonfailing tissue (p < 0.05). The time until maximal inotropic effect developed after ouabain was significantly shorter in NYHA IV (mean 150 +/- 16 min) than in nonfailing myocardium (mean 240 +/- 20 min). These results suggest that human failing myocardium exerts and enhanced sensitivity to cardiac glycosides and Na+ -channel activators, possibly because of enhanced expression of the Na+/Ca2+ exchanger or because of an altered intracellular Na+ -homeostasis.

Adolescent↗
PubMed1996

Detection of residual leukemic cells in acute lymphocytic leukemia of B-cells (B-ALL) by a nonradioactive PCR-based technique.

Patients suffering from B-ALL were analyzed for residual leukemic cells by means of the PCR technique. Using primers specific for conserved domains of each VH-family and a primer universal to the JH-regions, immunoglobulin gene fragments have been amplified from bone marrow aspirate-derived DNA in six cases of B-ALL. Cloning and sequencing of the amplified fragments revealed the usage of the VH251-family in two cases. VH-families 1 and 3 were used in one case each. One patient showed abnormal rearrangement with the participation of two JH regions. In the remaining case a VH6 germline fragment was involved in an apparently abnormal rearrangement. Based on the sequence information, clone-specific DIG-labeled probes were designed and used in subsequent nonradioactive hybridization protocols to estimate the prevalence of residual leukemic cells in the course of therapy. Five of the six cases of B-ALL could be included in the rest cell analysis. In all patients investigated, residual leukemic cells were detected, independent of whether a normal or apparently abnormal rearrangement has occurred. These findings imply that the prevalence of residual leukemic cells in B-ALL indicates a high risk of relapse.

Adult↗
PubMed1995

Quantification of aminopeptidase N mRNA in T cells by competitive PCR.

The aminopeptidase N (CD13, EC 3.4.11.2) is a well-characterized surface molecule expressed in a variety of cell types and species. Recent data indicate an expression of the APN mRNA and the corresponding aminopeptidase activity in human peripheral T cells and related cell lines as well. Here, the sensitive method of competitive PCR was used to quantify low amounts of APN mRNA in T cell lines. An APN cDNA fragment enshortened by a deletion of 87 bp was used as an internal APN-specific standard. The myelo-monocytic cell line U937 and the lymphoid T cell lines HuT78 and H9 contain 2.3 x 10(7), 5.9 x 10(6) and 5.6 x 10(6) copies/micrograms total RNA, corresponding to 160, 70 and 50 copies/cell, respectively. These data have been confirmed by determination of the APN activity, that represents a fraction only of the total cellular neutral aminopeptidase activity in hematopoetic cells. In the case of the CD13-positive cell line U937, approximately 60-70% of the total neutral aminopeptidase activity could be attributed to APN. In contrast, only a minor fraction (5-20%) of the cellular neutral aminopeptidase activity in the T cell lines H9 and HuT78 represents APN. The results suggest that APN gene expression within the hematopoetic system is not restricted to myelo-monocytic cells, instead a low APN expression may be a common feature of lymphocytes, at least of T cells, too.

Base Sequence↗
PubMed1994

Expression of the aminopeptidase N (CD13) gene in the human T cell lines HuT78 and H9.

The membrane-bound metalloprotease aminopeptidase N (APN, CD13, EC 3.4.11.2) is a well-established marker of normal and malignant cells of the myelomonocytic lineages. It is also expressed by leukemic blasts of a small group of patients suffering from acute or chronic lymphoid leukemia. CD13-specific monoclonal antibodies do not bind to the surface of normal B lymphocytes and APN-mRNA was not detectable by Northern analysis in normal lymphocytes or in T cell lines. A recent paper, however, describes the expression of CD13 on concanavalin A-stimulated T cells. Here, by means of enzymatic amplification of cDNA, cloning and sequencing of amplified cDNA fragments, in situ hybridization, and transcription start site mapping, evidence is provided that the human T cell lines HuT78 and H9 do contain APN-mRNA. Ala-pNA-hydrolyzing activity was detected in viable cells and cell homogenates as well as in size-fractionated protein fractions. Alanine-beta-naphthylamide (Ala-beta NA)-hydrolyzing activity was readily detectable after isoelectric focusing and blotting and was shown to be localized exclusively intracellularly in both cell lines. Usage of aminopeptidase-specific effectors revealed this activity to be distinct from CD13.

Aminopeptidases↗
PubMed1992

Monitoring of redox-state of respiratory enzymes and myoglobin oxygenation in the working rat heart in normoxia and oxygen deficiency.

The cellular oxygen supply in the isolated, hemoglobin-free perfused, working rat heart can be determined by measurements of myoglobin oxygenation. However, for a precise analysis of mitochondrial hypoxia and anoxia (pO2 < 0.01 Torr) redox-state of respiratory enzymes must be known. By use of the EMPHO (Frank et al. 1989) it is possible to perform a high speed spectrometry within very small tissue volumes. Because of the characteristic absorption spectra of oxygenated and deoxygenated myoglobin and of the oxidized and reduced cytochrome aa3 within the wavelength interval from 500 to 630 nm it is possible to isolate these two pigments from the remission spectra and to determine the oxygenation state of myoglobin and the redox-state of cytochrom aa3.

Animals↗
PubMed1992

[Validation of the Child Speech Test using the Heidelberg Speech Development Test with six-year-old preschool children].

We present and discuss results from researches to validate a new test for diagnosing speech-competences of preschool children KISTE. 41 children (6;0-6;11) were tested both with KISTE and HSET. Correlations, factor- and clusteranalysis showed that the structure of both tests is different and that KISTE has a good differential validity for both aspects of speech: communicative and structural.

Child↗
PubMed1991

Spectrometric measurements in the anterior eye vasculature of the albino rabbit--a study with the EMPHO I.

Intracapillary haemoglobin oxygenation (oxygen saturation) and haemoglobin concentration (corresponding with blood volume) were measured in different regions of the albino rabbit anterior eye segment using the Erlangen micro-lightguide spectrophotometer (EMPHO I). The tip of the microlightguide (active diameter 210 microns, cover diameter 0.5 mm) was placed in the vitreous by a goniometric mounting and controlled by a micromanipulator. After stepwise elevation of intraocular pressure (IOP) the haemoglobin oxygenation decreased slightly in the iris at an IOP-level of 60 mmHg, whereas in the three regions of the ciliary processes and in the peripheral choroid haemoglobin oxygenation did not decrease until values of 80 mmHg were applied. In contrast, haemoglobin concentration decreases when the IOP increases in all regions except in the pars plana where the haemoglobin concentration increased at 40 and 60 mmHg. The most pronounced reactive hyperaemia was found in the major ciliary processes. Quite passive changes of haemoglobin concentration were seen in the peripheral choroid. Effective regulatory mechanisms must exist which result in IOP-independent (20-60 mmHg) haemoglobin oxygenation, whereas pronounced changes in intracapillary haemoglobin concentration were observed. The decay of both parameters at IOP 80 mmHg is indicative of an exhaustion of their regulatory capacity. Epinephrine topically applied onto the conjunctival sac or injected into the common carotid artery led to a short-term decrease of haemoglobin concentration and, later, a hyperaemic response. A short-term haemoglobin deoxygenation was found especially in the major ciliary processes.

Animals↗