Search PubMed⌕ Search

Biomedical subjects

J Stein

Publications and source records attributed to J Stein.

At least 271 records · Page 15Linked to original sources

Preparation of basolateral membrane vesicles from rat enterocytes: influence of different gradient media.

An attempt was made to assess whether the choice of the gradient media could influence the yield of basolateral membrane vesicles isolated from the rat intestine as well as their functional characteristics. Crude membranes prepared in the same way were therefore centrifuged with 10% Percoll, on a discontinuous sucrose gradient or on a continuous sorbitol gradient. The protein yield was significantly higher with the Percoll gradient than with sucrose and sorbitol gradient centrifugation (2.7 +/- 1.0%; 0.4 +/- 0.1%; 0.6 +/- 0.2%, respectively). Enrichment in Na+,K+-ATPase was similar in all three preparations (8.50 +/- 2.34; 8.22 +/- 4.78; 8.20 +/- 2.08). However, contamination with brush border membranes was significantly higher after Percoll gradient centrifugation and negligible after the use of the other two gradient media. Transport of D-glucose in the BLM prepared by Percoll gradient centrifugation also indicated some contamination with functional brush-border membranes. An attempt to purify basolateral membrane vesicles after Percoll gradient centrifugation with Ca2+ precipitation, however, reduced the protein yield to less than 1%. We conclude that in the preparation of basolateral membrane vesicles from the rat enterocytes each of the gradient media may have certain advantages and disadvantages, which should be considered according to the purpose of the preparation.

Animals↗

Reduced postheparin plasma diamine oxidase activity in patients with chronic renal failure.

The intestinal mucosa plays a fundamental role as the site for absorption of nutrients, and as an important barrier against potentially harmful agents in the gastrointestinal lumen. The functional integrity of the small bowel in patients with chronic renal failure was measured by post-heparin-diaminoxidase (DAO) activity in a low and safe dose of heparin (5000 IU) in patients with chronic renal failure participating in a hemodialysis program. The results of the study provide one of the first observations about a diminished DAO-response in patients with chronic renal failure reflecting a mucosal abnormality in these patients which has to be further characterized. Our findings also show that this reduced DAO activity is not caused by the effect of uremic toxins. These results further show that a smaller and safer i.v. dose of heparin (5000 IU or less) will release DAO activity as reliably as high doses used by others. We therefore conclude that postheparin DAO activity might be used as a new noninvasive test reflecting intestinal function in patients with chronic renal failure.

Adult↗

Peripheral blood stem cells as a source for hemopoietic recovery following autologous bone marrow transplantation in childhood malignancy.

In an attempt to improve hemopoietic recovery after autologous bone marrow transplantation (ABMT), a project of peripheral blood stem cells (PBSC) harvest was initiated. Thirty-six children awaiting ABMT underwent 126 PBSC harvests primed by a conventional scheduled chemotherapy course and rGMCSF 5 micrograms/kg per day/s.c. Ages ranged from 12 months to 19 years and weight from 9 to 84 kg. PBSC harvest was carried out using the Fenwall CS 3000 Plus with the small volume collection chamber at a maximum whole blood flow rate of 15-45 ml/min; total volume processed was 1,700-10,000 ml. Total nucleated cells per collection was 0.35-5.62 x 10(8) cells per kg, and the number of CD34+ cells, 0.23-1.1 x 10(6)/kg. The number of colony forming units-granulocyte macrophages (CFU-GM) varied in these heavily pretreated patients from 0 to 5.3 CFU-GM x 10(4)/kg per collection. Immunophenotyping of the cells collected was performed by double staining for CD34, CD33, CD15, CD71, Ia and CD56. Most of the CD34+ cells were found to be CD38+; some were CD33+ and some CD33-. Low coexpression of CD34+ CD71+ cells may correlate with the low proliferating capacity of PBSC as compared to the BM cells. To date 22 children have undergone transplantation using combined autologous PBSC and bone marrow. We conclude that PBSC harvest is a feasible and safe procedure even in small children, and can be successfully performed following scheduled chemotherapy and administration of growth factors, resulting in substantial yield, also in heavily pretreated patients. This procedure is recommended in responding high risk patients at the stage of minimal residual disease and may replace ABMT in the future.

Adolescent↗

[Diagnosis of steatorrhea].

Several indirect parameters of steatorrhea and quantitative measures of fecal fat excretion are described and in use as diagnostic tools for the detection and the differential diagnosis of malabsorption syndromes. The knowledge of their rationals, principles and also of technical aspects is an important foundation for both their application and interpretation, and thereby enables a rational diagnostic approach on a pathophysiological base. This synopsis of traditional, present and innovative, emerging methods for the diagnosis of steatorrhea covers the advantages of the respective techniques (fecal weight, fecal aspect, microscopy [Sudan stain], steatocrit, serum beta-carotinoids, titrimetric [van de Kamer] and gravimetric analysis, 14CO2-breath tests, NMR-spectroscopy, near infrared spectrometric analysis [NIRA], and 13CO2 stable isotope breath tests for fat maldigestion and -absorption), prerequisites for their application, and practical recommendations for specific situations. It may thus serve as a practical guide to the spectrum of 'coprophilic' and 'coprophobic', of 'low budget"- and 'high tech'-methods for assessing steatorrhea.

Celiac Disease↗

Expression of the VLA beta 1 integrin family in bladder cancer.

Integrins are a family of transmembrane heterodimers, many of which function as receptors for extracellular matrix molecules and play a role in adherence to and motility on matrix components. Because of these functions, integrins are suspected of participating in metastatic processes. We investigated the expression of beta 1 integrins in human bladder cancer cell lines and tissues. Expression of beta 1 integrins on cultured bladder cancer cell lines was evaluated by flow cytometry, of 8 cell lines tested, alpha 1 was found in 4, alpha 2 and alpha 3 in all 8, alpha 4 in 1, and alpha 5 in 3. These results were in sharp contrast to the expression detected by immunostaining tissues containing normal urothelium and low stage (noninvasive) and high stage (invasive) bladder cancers. All normal urothelial tissues tested expressed alpha 2 and alpha 3 and none expressed alpha 1, alpha 4, or alpha 5. Similarly, a majority (77%) of low stage (noninvasive) bladder cancers stained positively for alpha 3, whereas only 6 of 13 expressed alpha 2 and none expressed alpha 1, alpha 4, or alpha 5. Among invasive bladder cancers, alpha 1 was detected in 7%, alpha 2 in 24%, alpha 3 in 68%, alpha 5 in 10%, and alpha 4 was not found in any samples. These results indicate that integrin expression in cultured human bladder cancer cell lines does not represent expression observed in tissue samples and may reflect adaption to or selection during tissue culture conditions. A progressive loss of alpha 2 expression is seen from normal urothelial cells through invasive bladder cancers. This loss may contribute to an invasive phenotype by a loss of the cell-cell adherence function mediated by the alpha 2 beta 1 and alpha 3 beta 1 integrins.

Humans↗

A synthetic peptide of the effector domain of rab3A stimulates inositol 1,4,5-trisphosphate production in digitonin-permeabilized pancreatic acini.

In this study we report that a synthetic peptide of the effector domain of rab3A (rab3AL(33-48)) stimulates both amylase secretion and inositol 1,4,5-trisphosphate (IP3)-accumulation in digitonin-permeabilized pancreatic acini in an analogous way to cholecystokinin-octapeptide (CCK8). Maximum CCK8-induced IP3-accumulation was observed at five seconds after addition of CCK8 to the acini. Maximum rab3AL(33-48)-induced IP3-production occurred 15 to 30 seconds after addition of rab3AL(33-48); then the acinar IP3 content declined towards the basal level. Heparin, an inhibitor of IP3 binding to its receptor, inhibited both rab3AL(33-48)- and CCK8-stimulated amylase secretion without affecting the response to vasoactive intestinal polypeptide. rab3AL(33-48) had no effect in intact acini, indicating that the site of action of rab3AL(33-48) is intracellular. We conclude that rab-like small molecular weight GTP-binding proteins regulate phospholipase C activity and thereby amylase secretion from inside of the cell.

Amylases↗

Bone metabolism before and after irradiation with ultraviolet light.

The beneficial effects of ultraviolet light on cutaneous vitamin D synthesis, calcium metabolism, and bone formation are well known. Regarding the increasing fear of side effects from ultraviolet B (UV-B), lamps with less energy in the UV-B range have been developed. Two spectra with differences in the emission of UV-B have therefore been evaluated for their influence on calcium metabolism. A group of 24 healthy male volunteers was subdivided into two treatment groups. Group 1 was exposed to lamps with higher energy of total UV-B but less energy at the wavelengths below 300 nm than the lamps used in group 2. All subjects were irradiated ten times within 12 days. Exposure time was 3 min in the first session and time of exposure was increased by 10% in every following irradiation (suberythematous doses only). Before the first irradiation, 3 days after the last exposure, and after 4 more weeks, the serum parameters of bone metabolism were determined by standard laboratory methods. Significantly increased levels of 25-hydroxyvitamin D3 were found in both groups. There was only a slight increase of 1,25-dihydroxyvitamin D3. Parathyroid hormone decreased significantly in group 2 only. The data would suggest beneficial effects on bone metabolism for both regimens. The observed effects were more pronounced when shorter wavelengths (group 2) were applied, although the total energy of UV-B was lower in these lamps.

Adult↗

A fast and efficient method for simultaneous X and Y in situ hybridization of human blastomeres.

PURPOSE: To use a 6-hr fluorescence in situ hybridization (FISH) procedure involving fluorochrome-labeled probes to determine the gender of blastomeres from arrested biopsied human embryos. RESULTS: Simultaneous detection of X and Y chromosomes was performed on 68 blastomeres with this technique. The FISH efficiency for gender determination was 95.5% (65/68). In addition, rehybridization with chromosome 8-specific probes was performed to determine the ploidy of blastomeres with more than two sex chromosomes. CONCLUSION: This technique offers an alternative to polymerase chain reaction for the preimplantation diagnosis of X-linked diseases and can also be used for ploidy assessment.

Biopsy↗

Effects of guanine nucleotides on bombesin-stimulated signal transduction in rat pancreatic acinar cells.

To study the role of guanine nucleotide binding proteins (G proteins) in bombesin receptor signal transduction, we investigated the effects of guanine nucleotide analogues and of the G protein activator NaF on bombesin-induced amylase release, inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) production and release of intracellular Ca2+ in rat pancreatic acini. In digitonin-permeabilized acini, guanosine 5'-[gamma-thio]triphosphate (GTP gamma S), a well-known activator of G proteins, potentiated bombesin-induced Ins(1,4,5)P3 production and increased amylase release at low bombesin concentrations (< 10 nM). By contrast, GTP gamma S decreased bombesin-stimulated amylase release at high bombesin concentrations (> 10 nM). Fluoride (10 mM), another G protein activator, had similar effects to GTP gamma S on amylase release. However, unlike GTP gamma S it had no effect on Ins(1,4,5)P3 production and release of intracellular Ca2+ induced by high bombesin concentrations. GDP and its analogues, such as 2'-desoxyguanosine 5'-diphosphate (dGDP) or guanosine 5'-[beta-thio]diphosphate (GDP beta S), inhibit activation of G proteins. GDP and dGDP both inhibited amylase release and Ins(1,4,5)P3 production at all bombesin concentrations tested. In contrast, GDP beta S mimicked the effects of GTP gamma S on bombesin-stimulated amylase release and Ins(1,4,5)P3 accumulation. In conclusion, we suggest that bombesin receptor-mediated signal transduction involves G proteins in pancreatic acini. The correlation between inhibition of maximum-stimulated enzyme secretion and further increase in Ins(1,4,5)P3 production in response to GTP gamma S at high bombesin concentrations suggests that overstimulation of phospholipase C inhibits amylase release. The discrepant effects of GDP and of GDP beta S on phospholipase C activity and amylase release might be due to the ability of GDP beta S, but not of GDP to activate G proteins persistently after phosphorylation by G protein-associated GDP kinases.

Amylases↗

Medial paraspinal muscle electromyography: techniques of examination.

We describe two techniques, developed through cadaveric dissections, for the electromyographic examination of the lumbar interspinal muscles and medial lumbar multifidi. Previous anatomic studies have indicated that these medial paraspinal muscles share unisegmental innervation by a single nerve root. Electromyography of these muscles may therefore improve the localization of lumbosacral radiculopathy. The ability of the two techniques to achieve appropriate needle placement was confirmed using percutaneous injection of colored latex into cadavers. The safety and feasibility of the techniques were confirmed in healthy volunteers. Further studies are underway in subjects with lumbosacral radiculopathy to determine the clinical utility of these techniques.

Adult↗

Sex distribution in arrested precompacted human embryos.

Evidence of sexual dimorphism before fetal gonadal differentiation in mammals has been accumulating, suggesting that male embryos develop faster than female ones. The current investigation was performed to evaluate whether the development rate of precompacted human embryos is controlled by sex chromosomes. Sex was determined by polymerase chain reaction and fluorescence in situ hybridisation in 172 arrested embryos derived from in vitro fertilisation. The sex ratio (1.02:0.98) did not differ significantly from 1:1. Although more males appeared to have greater fragmentation, the difference between the sex ratios of highly fragmented and normal embryos (1.08:0.92) was not significant. Arrested female embryos had a tendency to exhibit more than five nuclei and less than 10% fragmentation, but the trend was not statistically significant. The current results suggest that the first developmental block in human embryos occurs prior to and shortly after genomic activation and is not determined by the presence of the Y chromosome.

Blastomeres↗

Tumour necrosis factor alpha stimulates the activity of the human cytomegalovirus major immediate early enhancer/promoter in immature monocytic cells.

Both tumour necrosis factor alpha (TNF-alpha) and phorbol 12-myristate 13-acetate (PMA) stimulated human cytomegalovirus (HCMV) major immediate early (IE) enhancer/promoter activity in the HL-60 granulocyte/monocyte progenitor cell line when added to transfected cells. In U-937 monocytic cells, by contrast, TNF-alpha had no stimulatory effect and the addition of PMA produced only marginal stimulation. In the mature THP-1 monocytic cell line and in differentiated HL-60 cells, addition of TNF-alpha caused inhibition of the IE enhancer/promoter activity. The stimulating effect of PMA, as observed in the other cell lines, however, remained. Thus the effect of TNF-alpha on the major IE enhancer/promoter activity is determined by the degree of differentiation of the infected cells. Unlike TNF-alpha and PMA, the interleukins IL-1, IL-3, IL-6 as well as the cytokine GM-CSF were found to have no detectable influence on the activity of the IE enhancer/promoter activity which, likewise, was not affected by the presence of the modulator sequence. Since premonocytic cells are suggested to be sites of HCMV latency, the stimulation by TNF-alpha could be of potential pathophysiological significance.

Cell Line↗

The stability of binocular fixation during reading in adults and children.

The authors recorded the binocular eye-movements of children and adults while they read single words appropriate for their reading ability. Normal data were obtained from nine- to 11-year-old primary-school children and adults, because very little is known about changes in binocular vergence during reading. These normal data were compared with those from poor readers of the same age who had passed or failed the Dunlop Test. On average, normal children made larger vergence errors while they fixated words than did adults. There were no differences between the groups of children, therefore the authors conclude that poor vergence control during reading fixations is not the immediate cause of the non-word error effect found among children who fail the Dunlop Test.

Adult↗