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Biomedical subjects

M Fromm

Publications and source records attributed to M Fromm.

At least 127 records · Page 7Linked to original sources
PubMed1987

Antineoplastic activity of conjugates of lipids and 1-beta-D-arabinofuranosylcytosine.

Five different lipid conjugates of 1-beta-D-arabinofuranosylcytosine (ARA-C) were tested in comparison with ARA-C, the ether lipid ET-18-OCH3 (1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine) and their equimolar mixtures. The compounds were tested in vitro for cytotoxicity in the trypan blue dye exclusion test with cells from six different leukemias, one glioblastoma and two bronchogenic carcinomas of human origin. The compounds were given in vivo to assess their therapeutic activity against 3-Lewis lung carcinoma (3-LL) of syngeneic C57Bl6 mice. Although some of the conjugates have shown cytotoxic activity in vitro against the cell samples tested, they have not revealed higher cytotoxicity than ET-18-OCH3, ARA-C or their equimolar mixtures. In these experiments, ARA-CDP-D,L-MBA was the conjugates significantly inhibited tumor growth and also increased survival of C57Bl6 mice with intraperitoneally (ip) implanted 3-LL. In these experiments, ARA-CDP-D,L-PTBA, ARA-CDP-D,L-PBA, ARA-CDP-L-dipalmitin and ARA-CDP-D,L-PCA were more active than either the parent compounds ARA-C and ET-18-OCH3 alone or their equimolar mixtures. Furthermore, when the conjugates were injected as adjuvant chemotherapy shortly after the surgical removal of the primary 3-LL, they inhibited the metastasis of 3-LL to the lungs of the animals, demonstrated by an increase of the survival time and the number of surviving animals. The mode of action of these new antineoplastic compounds still is speculative.

Animals↗
PubMed1986

Late effects after treatment of twenty children with soft tissue sarcomas of the head and neck. Experience at a single institution with a review of the literature.

Twenty children with soft tissue sarcomas of the head and neck, treated at the Children's Hospital of Philadelphia and the Hospital of the University of Pennsylvania from 1972 to 1981, were evaluated for the late deleterious effects of treatment. All patients received radiation therapy and combination chemotherapy with vincristine, dactinomycin, and cyclophosphamide; certain patients also received Adriamycin (doxorubicin). All had ophthalmologic, otologic, growth, and cosmetic evaluations; 15 also had dental and maxillofacial examinations. The median age at diagnosis was 6 years (range, 7 months-13 years). Median follow-up from time of diagnosis was 5.5 years with a minimum of 3 years in all but four patients. The major problems encountered were related to the eyes (xerophthalmia and cataracts), ears (hearing loss), teeth (maleruption and caries), glandular structures (xerostomia, hypopituitarism), and development (craniofacial deformity). It is concluded that children treated for soft tissue sarcomas of the head and neck with combined modality therapy, including radiation enhancers, may show a variety of late treatment-related adversities. These children require close multidisciplinary follow-up for detection of late effects in order that appropriate prophylactic or symptomatic treatment can be instituted to minimize their consequences.

Adolescent↗
PubMed1986

Epithelial and subepithelial resistance of rat large intestine: segmental differences, effect of stripping, time course, and action of aldosterone.

Epithelial and subepithelial electrical resistances of rat large intestine were measured by means of a 4-electrode AC impedance technique in three segments, colon ascendens, colon descendens and rectum. Epithelial resistance of colon ascendens and colon descendens was about 35 omega X cm2 and not different between these two segments. It was, however, about 3 times higher in rectum (99 omega X cm2). This finding is in accord with our previous observation of about 3-fold higher net fluxes of ions and water in colon ascendens and colon descendens than in rectum. It confirms the concept of a main functional difference between the terminal part of the large intestine (rectum) and the more proximal segments (colon). The acutely (within hours) varied level of aldosterone by keeping the rats for 7 h in anaesthesia caused in the rectum a more than 10-fold increase in short circuit current (Isc) and transepithelial voltage but no significant decrease in resistance. Similarly, the decline in Isc, as regularly observed in the early phase of in vitro measurements on partially stripped large intestine, was paralleled by voltage changes but not by changes in resistance. We conclude that the wide range of resistance values published so far was caused to a great extent by including various portions of colon or rectum. By comparing intact (not stripped) and partially stripped preparations (muscularis propria removed) of the rectum it was shown that partial stripping did not alter the epithelial resistance but reduced the subepithelial resistance in this segment from 26 to 8 omega X cm2, or by 68%.(ABSTRACT TRUNCATED AT 250 WORDS)

Aldosterone↗
PubMed1985

Epithelial and subepithelial contributions to transmural electrical resistance of intact rat jejunum, in vitro.

Epithelial and subepithelial resistance of rat jejunum was measured in vitro by two independent methods. (i) Transepithelial AC impedance data were interpreted in terms of a simple parallel RpCp element (representing the epithelial cell layer) in series with an ohmic resistor RS (representing the subepithelial layers). (ii) In separate experiments, the tip of a microelectrode was positioned between epithelium and subepithelial layers and the respective resistances were obtained from DC-pulse voltage divider ratios between both structures. The total tissue resistance as measured in conventional Ussing-chamber experiments (49 +/- 4 Ohm X cm2, mean of both methods) was formed to 81 +/- 6% (40 +/- 3 Ohm X cm2) by subepithelial layers and to only 19 +/- 3% (9 +/- 1 Ohm X cm2) by the epithelial cell line. We conclude that rat jejunum is more conductive than assumed so far. In in vitro flux studies on intact jejunal sheets a pronounced back-diffusion of absorbed substances will lead to an underestimation of the true net transport capacity of this structure. This error averages about fivefold and will be found likewise in conventional short-circuit measurements.

Animals↗
PubMed1985

Protamine reversibly decreases paracellular cation permeability in Necturus gallbladder.

Protamine, a naturally occurring arginine-rich polycationic protein (pI 9.7 to 12), was tested in Necturus gallbladder using a transepithelial AC-impedance technique. Protamine sulfate or hydrochloride (100 micrograms/ml = 20 microM), dissolved in the mucosal bath, increased transepithelial resistance by 89% without affecting the resistance of subepithelial layers. At the same time, transepithelial voltage (psi ms) turned from slightly mucosa-positive values to mucosa-negative values of approximately +1 to -5 mV. The effect of protamine on transepithelial resistance was minimal at concentrations below 5 micrograms/ml but a maximum response was achieved between 10 and 20 micrograms/ml. Resistance started to increase within 1 min and was maximal after 10 min. These effects were not inhibited by serosal ouabain (5 X 10(-4) M) but could be readily reversed by mucosal heparin. The sequence of protamine effect and heparin reversal could be repeated several times in the same gallbladder. Mucosal heparin, a strong negatively charged mucopolysaccharide, or serosal protamine were without effect. Mucosal protamine reversibly decreased the partial ionic conductance of K and Na by a factor of 3, but did not affect Cl conductance. Net water transport from mucosa to serosa was reversibly increased by 60% by protamine. We conclude that protamine reversibly decreases the conductance of the cation-selective pathway through the tight junction. Although this effect is similar to that reported for 2,4,6-triamino-pyrimidinium (TAP), the mechanism of action may differ. We propose that protamine binds to the apical cell membrane and induces a series of intracellular events which leads to a conformational alteration of the tight junction structure resulting in decreased cationic permeability.

Animals↗
PubMed1985

Expression of genes transferred into monocot and dicot plant cells by electroporation.

We have developed a general method for electrically introducing DNA into plant cells. Gene transfer occurs when a high-voltage electric pulse is applied to a solution containing protoplasts and DNA. Carrot protoplasts were used as a model system to optimize gene-transfer efficiency, which was measured 24-48 hr after electroporation by the amount of chloramphenicol acetyltransferase activity resulting from the expression of the introduced chimeric plasmids. Gene-transfer efficiency increased with the DNA concentration and was affected by the amplitude and duration of the electric pulse as well as by the composition of the electroporation medium. Our optimized gene-transfer conditions were effective when applied to tobacco and maize protoplasts, demonstrating that the method is applicable to both monocot and dicot protoplasts.

Acetyltransferases↗
PubMed1984

Complex regulation of simian virus 40 early-region transcription from different overlapping promoters.

During simian virus 40 lytic infection there is a shift in initiation sites used to transcribe the early region, which encodes large T and small t antigens. Early in infection, transcription is initiated almost exclusively from sites that are downstream of the origin of DNA replication, whereas transcripts produced later are initiated mainly from sites on the upstream side. We have used mutant virus and specially constructed plasmid DNAs to investigate the factors regulating this transcriptional shift. In our studies simian virus 40 large T antigen appears to mediate the shift in transcription in two ways: first, T antigen represses transcription at the downstream sites late in infection by binding to the region where these RNAs are initiated; second, T antigen promotes transcription from sites on the upstream side by its ability to initiate replication or amplification, or both, of the template DNA. In addition, transcription from the downstream sites is heavily dependent on enhancer sequences located in the 72-base-pair repeat region, whereas transcription from the upstream sites late in infection does not require enhancer sequences. Thus, different overlapping promoters regulate simian virus 40 early-region expression in a manner that apparently coordinates the production of large T antigen with the increase in viral DNA.

Animals↗
PubMed1983

Time course of aldosterone and corticosterone plasma levels in rats during general anaesthesia and abdominal surgery.

The transepithelial voltage (psi ms) of rat rectum in vivo increases for several hours in experiments under general anaesthesia. So far this was attributed by indirect evidence to increasing aldosterone plasma levels during the course of the experiment. We performed direct measurements of aldosterone and corticosterone plasma concentrations during intestinal perfusion experiments on barbiturate anaesthetized rats. Experiments were terminated for blood sampling at 10, 75, 300, 400, 800, or 1,800 min, respectively. (i) After 75 min of anaesthesia, surgical preparation was finished and plasma levels of aldosterone and of corticosterone were found increased by the factors 5 and 3, respectively, as compared to conscious controls. (ii) During the following 12 h, aldosterone further increased to levels 10 times as high as those of controls. In contrast, during the same period corticosterone slowly decreased but still remained elevated as compared to controls. (iii) The increase of both hormones was attenuated when abdominal surgery was omitted. (iv) The use of pentobarbital (Nembutal) instead of thiobarbital (Inactin) did not influence the adrenal response. (v) In adrenalectomized rats a continuous substitution with 65 ng X h-1 X kg-1 BWT aldosterone resulted in plasma levels as high as in conscious intact animals. (vi) Rectal psi ms started to move to higher lumen-negative values with a time delay of 1-1 1/2 h as compared with the increase of hormone levels. psi ms then stayed elevated until to the end of the experiments. We conclude that in vivo experiments of several hours duration in thio- or pentobarbital anaesthetized rats take place under conditions of aldosterone and corticosterone plasma levels which are high as compared to those of conscious unstressed animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Abdomen↗
PubMed1983

Simian virus 40 early- and late-region promoter functions are enhanced by the 72-base-pair repeat inserted at distant locations and inverted orientations.

Tandemly repeated 72-base-pair (bp) segments located between nucleotides 107 and 250 of the simian virus 40 genome are essential for early region transcription. The functional requirement for the 72-bp repeat was supplied even when that segment was translocated to several locations distant from, and in different orientation, relative to, the promoter. Regardless of the position of the 72-bp enhancer segment, transcription was initiated at the same locations as with the normal promoter. Translocation of the 72-bp repeat segment to other sites in the genome resulted in the appearance of DNase I hypersensitivity at that site in the intranuclear viral minichromosomes. One of the translocations which did not produce enhancement of early- and late-region expression also failed to create a DNase I-hypersensitive site at the translocated 72-bp segment.

Base Sequence↗
PubMed1983

Cytotoxicity of thioether-lysophospholipids in leukemias and tumors of human origin.

Thioether-lysophospholipids inhibited the in vitro incorporation of [3H]thymidine into blasts of 8 leukemias, lymphocytes of 3 chronic lymphocytic leukemias, and cells of 12 different solid tumors of human origin. This effect correlated with trypan blue dye exclusion, which was used to assess cell damage. Scanning electron microscopy revealed severe membrane destruction after incubation with thioether-lysophospholipids. Cytostatic and cytotoxic effects of thioether-lysophospholipids were dependent on dosage and incubation time. Destruction of leukemic blasts was completed with greater than or equal to 5 micrograms/ml after an incubation of greater than or equal to 48 hr, but 10 to 20 micrograms/ml were necessary in solid tumors. Ester-linked 2-lysophosphatidylcholine was ineffective in the same dose range, which points to the requirement of the alkyl moiety in SN1 of the molecule for the antineoplastic properties of lysophospholipid analogues.

Adenocarcinoma↗
PubMed1983

Transcription in vivo from SV40 early promoter deletion mutants without repression by large T antigen.

Transfection of monkey cells with recombinant plasmids containing a beta-globin coding region fused to wild-type or deleted simian virus 40 (SV40) early region promoters has allowed an analysis of the transcriptional activity of these promoters in the absence of repression by large T antigen. We find that deletion of the TATA sequence does not reduce the transcriptional effectiveness of the promoter; however, the 5' ends of the transcripts are heterogeneous rather than being restricted to the usual sites. The short GC-rich repeat sequences between nucleotides 37 and 107 and the tandemly repeated 72-bp segment between nucleotides 107 and 250 are each essential for promoter function. The GC-rich repeats are functionally redundant and probably interchangeable, since several subsets of two or three of the GC-rich segments are sufficient. One copy of the 72-bp sequence is sufficient to permit transcription. Moreover, the 72-bp repeat sequences function normally even if they are inverted relative to their normal orientation.

Antigens, Viral↗
PubMed1982

Crypts are the site of intestinal fluid and electrolyte secretion.

The site of adenosine 3',5'-monophosphate-mediated fluid and electrolyte secretion across mammalian large intestine was found to be the crypts of Lieberkühn by means of two techniques. First, the formation of fluid droplets was visualized on the oil-covered mucosal surface directly over crypt duct openings when secretion was stimulated. Second, microelectrode impalement of individual surface and crypt cells revealed that only crypts cells produced a pattern of secretagogue induced alterations in membrane potential and resistance that was characteristic of secretory epithelia.

Amiloride↗
PubMed1982

Deletion mapping of DNA regions required for SV40 early region promoter function in vivo.

The SV40 early region promoter, previously localized to the DNA segment bounded by the HpaII and HindIII restriction sites (nucleotides 346 and 5171), was further defined by construction of an extensive set of deletions within this region and measurement of their effects on (a) viral DNA replication, (b) virus multiplication and the ability to complement early and late mutations, (c) transformation of rat cells, (d) large T antigen formation, and (e) the location of the 5' ends of early mRNAs. One set of mutations is represented by deletions that begin at the HpaII site and extend unidirectionally for varying lengths toward the BglI site at ori. A second set of mutants contains deletions that start at ori and extend unidirectionally for varying lengths towards the HpaII site. A third set of mutants, with deletions or duplications of various lengths and boundaries, lie between the HpaII and BglI sites. Our studies indicate the following. (a) Ori, the sequence needed for initiating SV40 DNA replication, extends from the sequences needed for initiating SV40 DNA replication, extends from the sequences needed to bind T antigen to the palindrome in site II to nucleotide 34, the late region edge of the AT block. Flanking sequences adjacent to the AT block facilitate DNA replication. (b) The SV40 early region promoter comprises two functionally distinct nucleotide sequence elements. One is flanked by nucleotides 5231 and 107, and contains the RNA initiation sites at nucleotides 5231-5237, a positioning element resembling the TATAAATA consensus sequence about 20-25 nucleotides upstream, and an RNA polymerase II recognition sequence contributed by short GC-rich sequences clustered between nucleotides 35 and 107; we refer to this as the RNA polymerase II interaction site. The second distinct sequence element is contained within each of two 72-bp segments located between nucleotides 107 and 250; the behavior of this element suggests that it may influence the accessibility of RNA polymerase II for the interaction site or the efficiency of RNA chain initiation. Large T antigen binding sites I, II, and III overlap with the putative RNA polymerase II interaction site; since large T antigen does not prevent elongation of RNA transcripts initiated upstream, T antigen probably represses early region expression by preventing RNA polymerase II binding to the promoter.

Chromosome Deletion↗
PubMed1981

Potassium transport across rabbit descending colon in vitro: evidence for single-file diffusion through a paracellular pathway.

The results of previous studies indicate that the bidirectional fluxes of K across short-circuited rabbit descending colon are attributable to passive diffusion through paracellular pathways and that this route is ten times more permeable to K than to Na and Cl. However, transepithelial diffusion potentials in the presence of large transepithelial Na and K concentration differences are much lower than those predicted by the "constant field equation" and appear to be inconsistent with this high K selectivity. The results of the present studies, designed to resolve this apparent contradiction, indicate that: (a) The ratios of the bidirectional transepithelial fluxes of K determined over a wide range of combined chemical and electrical potential differences conform reasonably well with those predicted by the Ussing flux-ratio equation. (b) The permeability coefficient of K (PK), determined from the net fluxes in the presence of concentration differences and from unidirectional fluxes under short-circuit conditions, decreases with increasing K concentration; in the presence of low K concentrations, PK is approximately ten-times PNa, but it approaches PNa in the presence of high K concentrations. PNa is not affected under these conditions. These results provide an explanation for the failure to observe large transepithelial diffusion potentials in the presence of large transepithelial Na and K concentration differences. In addition, these results are consistent with the notion that K diffuses across this preparation through two parallel pathways, one that does not discriminate among K, Na and Cl (a "free-solution" shunt) and another that is highly K selective and involves an interaction with one, or at most two, sites along the route.

Animals↗
PubMed1981

Some properties of KCl-filled microelectrodes: correlation of potassium "leakage" with tip resistance.

This study was undertaken in order to determine directly the rates of K leakage (JK) out of the tips of microelectrodes into a solution of 100 mM KCl (approximating the K concentration of the cell interior) and to relate these rates to the concentration of the filling solution and the tip resistance. The values of JK for electrodes filled with 3 M KCl having resistances of 16 and 30 M omega (when measured in 3 M KCl) were 10 and 5.5 fmol/sec, respectively. When the same electrodes were filled with 0.5 M KCl, the resistances (measured in 0.5 M KCl) increased to 62 and 115 M omega, respectively, and JK fell to 1.8 and 1.0 fmol/sec, respectively. These values are in reasonable agreement with what would be expected from theoretical considerations if leakage of KCl were the result of diffusion plus convective flow due to the hydrostatic pressure of the filling solution. We conclude that K leakage out of microelectrodes filled with 3 M KCl is unnecessarily high; leakage can be reduced fivefold by filling electrodes with 0.5 M KCl without incurring significant increases in tip or diffusion potentials or unmanageable tip resistances. Finally, the lowest rate of K leakage observed (1 fmol/sec) is still very considerable for the case of animal cells with an intracellular volume of approximately 1 pl and a K content of approximately 100 fmol. The finding of stable intracellular potentials, often for many minutes, in some tissues suggests that K which enters the cell rapidly diffuses into neighboring cells via high conductance intercellular communications.

Diffusion↗
PubMed1980

Versatile piezoelectric driver for cell puncture.

A simple and versatile tool facilitating micropuncture of small cells is described which utilizes a commercial piezoelectric element made from a stacked column of monomorph ceramic discs. The device is able to advance complete input stage-electrode-assemblies with high speed and can be used in combination with conventional micromanipulators. Advancing characteristics as recorded optically at high magnification demonstrated less axial vibration, although faster action, than two other modern micropositioners driven by step motors. In biological experiments on selected tissues (Necturus gallbladder epithelium, Amphiuma renal distal tubule cells, rabbit and human corneal endothelium) the combined use of micromanipulator and piezo-stepper was, in all cases, superior to the use of a micromanipulator alone: the percentage of successful cell penetrations increased, cell potentials were stable for a longer time, and the durability of electrode-tips improved.

Animals↗
PubMed1978

Segmental heterogeneity of epithelial transport in rat large intestine.

Functionally isolated segments of rat colon and rectum were perfused in situ in a closed loop system. Rectum was defined as the lower 25--35% of the length of large intestine (cecum excluded). Perfusion conditions were optimized at 0.5 ml.min-1 and 3 cm H2O luminal pressure. Variation of perfusion rate between 0.2 and 2 ml.min-1 did not influence net volume transport (JNV). Luminal distension following elevation of hydrostatic pressure to 18 cm H2O reversibly increased Jnv. Under control conditions Jnv and Na+-transport rates (JnNa) of colon were 2--3 times higher than those of rectum. In colon transepithelial electrical potential difference (psims) was time independent --12 mV (lumen negative) whereas rectal psims increased with time from --6 mV, reaching a plateau of --67 mV within 6 h. Amiloride 10(-4) mol.l-1 had no effect on psims, Jnv, and JnNa in colon but did slightly depress K+-secretion in colon descendens. In contrast, psims in rectum was dose-dependently depressed, being reversed to +7 mV at 10(-4) mol.l-1. Jnv and JnNa were decreased by half. Acetazolamide in addition to amiloride lowered the positive post-amiloride rectal psims by half. Adrenalectomy had no effect on colonic psims, but abolished psims of the rectum. A single dose of 40 microgram.kg-1 b.w. aldosterone during the experiment restored the typical time course of rectal psims, but did not affect psims in colon. It is concluded that aldosterone induces an amiloride-sensitive Na+-pathway only in rectum, but not in colon, and that colon and rectum differ basically in their transport properties, quantitatively as well as qualitatively, as do the kidney distal convoluted tubule and the cortical collecting duct.

Acetazolamide↗