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Biomedical subjects

K Eckert

Publications and source records attributed to K Eckert.

At least 37 records · Page 2Linked to original sources
PubMed1997

Evaluation of a new commercial microimmunofluorescence test for detection of antibodies to Chlamydia pneumoniae, Chlamydia trachomatis, and Chlamydia psittaci.

A new commercial test for chlamydial serology, the MRL-Micro-Immunofluorescent Test (MRL; MRL Diagnostics, USA) was compared with the standard microimmunofluorescence test (MIF) using sera from 246 patients. Chlamydia pneumoniae immunoglobulin G (IgG) antibodies were detected in 46.3% (MIF) and 64.2% (MRL) of sera and Chlamydia trachomatis IgG in 23.2% (MIF) and 25.2% (MRL); Chlamydia psittaci IgG antibodies were found with the MRL in 1% of the sera from a general population and in 17.3% of preselected sera with elevated complement fixation titers. Titers were usually higher with the MRL. IgG titers of > or = 1:512 were detected in only 2% of sera using the standard MIF but in 30% using the MRL. In 16 sera from three Chlamydia pneumoniae culture-positive patients, the diagnosis of acute infection could be confirmed serologically in one with the MRL test but in none with the MIF test, indicating a higher sensitivity of the MRL.

Antibodies, Bacterial↗
PubMed1997

Prothymosin alpha 1 effects, in vitro, on the antitumor activity and cytokine production of blood monocytes from colorectal tumor patients.

Recent animal studies demonstrate that prothymosin alpha 1 (ProT alpha) enhances the antitumor response by stimulation of mononuclear phagocyte functions. The present study was aimed at characterizing the in vitro effects by ProT alpha on blood monocytes from human colon cancer patients. Purified peripheral blood monocytes were studied in terms of tumor cytostatic ability and cytokine production after incubation with ProT alpha or interferon (rIFN-gamma) and transforming growth factor-beta (TGF beta), used as reference substances. SW620 colon carcinoma cells were used as tumor target cells in growth inhibition experiments. The level of baseline growth inhibitory activity of unstimulated patient's monocytes was significantly lower than that of normal monocytes. The defective antitumor activity of patient monocytes was associated with a higher production of the inhibitory monokines prostaglandin E2 (PGE2) and TGF beta. The stimulation of monocytes by ProT alpha and/or rIFN-gamma elevated the average antitumor activity in all donor groups. The ProT alpha-induced increase was associated with a significantly higher monocytic secretion of IL-1 beta and TNF-alpha. Moreover, the concentrations of TGF beta and PGE2 in the culture supernatants decreased significantly, when patient's monocytes were treated with ProT alpha and/or rIFN-gamma. Additionally, ProT alpha enhanced the diminished antitumor activity of TGF beta-treated normal monocytes. These results suggest that ProT alpha selectively regulates distinct functions of blood monocytes, the effect of this cytokine varying with the parameter and donor population examined. These data provide a rational and biological endpoint for further studies with ProT alpha as an activator of mononuclear function in colon cancer.

Aged↗
PubMed1997

Prothymosin alpha 1 effects in vitro on chemotaxis, cytotoxicity and oxidative response of neutrophils from melanoma, colorectal and breast tumor patients.

Immunoregulatory effects of thymic peptides on functions of polymorphonuclear leukocytes (PMNs) are poorly investigated. We studied the effects of prothymosin alpha 1 (Pro alpha 1) on PMNs from patients with colorectal tumors, breast tumors and melanoma (total n = 37) in comparison with healthy donors (n = 18), with respect to chemotaxis, cytotoxicity against HCT-116 colon tumor cells, oxidative response (chemiluminescence reaction) as well as expression of surface marker molecules. We found that Pro alpha 1 was equally effective in stimulating the chemotactic activity of PMNs from tumor patients and healthy donors (43% increase). PMNs from tumor patients, especially with breast tumor, showed a significant enhancement of cytotoxicity against the tumor target cells in comparison with healthy donors. With respect to the PMNs cytotoxicity, only about 50% of the colorectal tumor patients and healthy donors responded to Pro alpha 1 and FMLP. As to the oxidative response of PMNs, elevated levels were found only among colorectal tumor patients. Pro alpha 1 significantly increased the oxidative response in breast and colorectal tumor patients by 55% and 25%, respectively. Pro alpha 1 decreased the expression of CD16 on PMNs of healthy donors, but not that of CD11a, CD11b, CD11c, CD13, CD14, CD15 and CD32. Therefore, we suggest, that Pro alpha 1 may improve some PMN functions of tumor patients, associated with the proposed role in host-tumor interaction.

Aged↗
PubMed1997

Prothymosin alpha1 effects on IL-2-induced expression of LFA-1 on lymphocytes and their adhesion to human umbilical vein endothelial cells.

Prothymosin alpha1 (Pro alpha1) is known to stimulate in vitro and in vivo natural killer (NK) and lymphokine (IL-2)-activated killer (LAK) cells against tumor cells. In this process, LAK cells first adhere to endothelial cells in vivo, raising the question whether Pro alpha1 affects this interaction as well. The binding ability of peripheral blood lymphocytes (PBL) to human umbilical vein endothelial cells (HUVEC) was increased by incubation with IL-2 in a concentration-dependent manner, reaching a maximal value at 20U/ml IL-2. Although Pro alpha1 alone was without any stimulating effect, it significantly increased PBL binding to unstimulated HUVECs in combination with suboptimal IL-2 (5 and 10 U/ml). The combination of Pro alpha1 (1 microg/ml) and 5 U/ml or 10 U/ml IL-2 is as effective as 10 U/ml or 20 U/ml IL-2 alone. This Pro alpha1 effect on IL-2-activated lymphocytes was found to be augmented on IL-1 or tumor necrosis factor (TNF)-alpha-activated endothelial cells. Analyzing the effect of Pro alpha1 on IL-2-activated lymphocytes by flow cytometry revealed an increase of CD16, CD56, and CD18 surface marker expression, whereas CD3, CD11a/b, CD49d, and CD54 were not affected. In conclusion, Pro alpha1 functions as a mediator of the adhesion of IL-2-activated lymphocytes to HUVECs.

Antigens, CD↗
PubMed1996

A colorimetric immunoassay for the detection of E-cadherin and carcinoembryonic antigen (CEA) expression on human colon carcinoma cell lines in vitro.

An immunohistochemical assay, using 96-well microtiter plates and based on the avidin-biotin peroxidase reaction, was developed for the quantitation of E-cadherin and CEA expression on HT-29 and SW620 colon carcinoma cells. The optical density of the generated dye was measured after solubilization with alkaline SDS solution. The staining procedure was evaluated with respect to reproducibility (coefficient of variation and intra-class correlation coefficient). On HT-29 cells the level of agreement for both E-cadherin and CEA were substantial, on SW620 cells almost perfect. The method allows testing compounds for their differentiation inducing capacity in screening programmes on the basis of protein marker expression.

Cadherins↗
PubMed1996

Differences in bone mineral in young Asian and Caucasian Americans may reflect differences in bone size.

Bone mineral content (BMC) and areal bone mineral density (BMD) have been reported to be lower in Asian than in Caucasian adults. To determine if racial differences in bone mass are present in younger subjects and whether they reflect differences in estimated volumetric bone density or in bone size, we compared measurements of bone mineral in healthy young Asian- and Caucasian-American males and females. Bone mineral was measured at the lumbar spine (L2-L4), femoral neck (FN), and whole body (WB) by dual-energy X-ray absorptiometry (DXA) in 99 Asians (49 females, 50 males) and 103 Caucasians (54 females, 49 males) ages 9-26 years. Results were expressed as BMC, BMD, and apparent density (BMAD), an estimate of volumetric bone density that reduces the effect of bone size. Subjects were compared on the basis of chronological age as well as by Tanner stage to correct for potential differences in the timing of puberty. Habitual dietary intake and physical activity were also assessed and correlated with bone mineral. The Asian and Caucasian cohorts differed in body size, diet, and physical activity. Asian females were shorter than the Caucasian females at all stages of puberty and weighed less at pre-/early puberty (p < 0.05). Asian males were older than Caucasians at midpuberty (p < 0.01) and weighed less than the Caucasian males at pubertal maturity (p = 0.001). Asian youths also consumed less calcium and reported less weight-bearing activity. Racial differences were most apparent when comparing BMC data. Asian males had greater spine BMC at midpuberty and lower WB BMC at maturity (p < 0.05). Asian females had lower FN BMC through midpuberty and lower WB BMC in pre-/early puberty (p < 0.05). WB BMD and WB BMC/height values were significantly lower in mature Asian versus Caucasian males. No significant racial differences in BMAD were observed. Multivariate regression analysis indicated that the differences in BMD and BMAD between Asian and Caucasian subjects were largely attributable to differences in weight and pubertal stage, and, at the FN, in weight-bearing activity. Further, the explanatory variables were less strongly associated with BMAD than with BMD. In summary, no significant differences in BMD were found between Asian and Caucasian youths through midpuberty; however, WB BMD and WB BMC/height values were lower in Asian males at sexual maturity. We conclude that observed differences in bone mineral between Asians and Caucasians may be partially attributed to the smaller bone size of Asians.

Absorptiometry, Photon↗
PubMed1996

A rapid and sensitive fluorometric screening assay using YO-PRO-1 to quantify tumour cell invasion through Matrigel.

A new quantitative assay for the study of tumour cell invasion in vitro is described. Employing the novel fluorescent dye YO-PRO-1, cells that penetrate Matrigel-coated transwells are counted on the basis of dye-bound cellular nucleic acid content. Following transmigration, the cells in the lower compartments are lysed by freezing in water. After a brief incubation with YO-PRO-1, nucleic acid or DNA content is measured as fluorescence intensity in 96-well microplates and quantitated by a cell- or DNA-calibration curve. Using standard curves, a linear relationship between fluorescence intensity and cell number was found in the range tested (from 100 to 80 000 cells). The mean relative intra- and inter-assay variability of the cell quantitation in this range was 3.5 and 4.2%, respectively. When applied to Matrigel invasion studies, as few as 400 cells could be counted. The quantitation could be performed within 3 h. HCT 116, MDA MB 231 and HT 29 cells were investigated as examples of tumour cells with different invasive abilities in the 48-h Matrigel invasion assay. Using YO-PRO-1, 6.5 +/- 0.6% invasive HCT 116 cells and 52.6 +/- 4.5% MDA MB 231 cells (percentage of the inoculated cell population) were measured. HT 29 cells were practically non-invasive. These results were confirmed by visual scoring of DAPI-stained nuclei. In conclusion, the main advantages of the assay are its sensitive, reproducible and rapid quantitation of tumour cell invasion in vitro and the applicability to extended sample numbers by measuring in 96-well microplates.

Basement Membrane↗
PubMed1996

Reversible cardiomyopathy in a child with Addison's disease.

An 11-year-old boy in acute adrenal crisis was subsequently diagnosed with Addison's disease and presumably had an unrecognized preceding chronic adrenal insufficiency. His adrenal insufficiency and hypovolemic shock were promptly diagnosed and treated in the emergency room. However, 24 h later, signs of congestive heart failure developed and an echocardiogram revealed cardiomyopathy. He responded to i.v. dopamine and his cardiomyopathy resolved over some days. Acute adrenal insufficiency and actual ventricular dysfunction had not been described in humans until last year; this is the second reported case of cardiomyopathy in a patient with acute adrenal insufficiency.

Acute Disease↗
PubMed1995

Prothymosin alpha: a biologically active protein with random coil conformation.

Prothymosin is an acidic protein with an unusual amino acid composition. Though its exact function is not yet known, its high evolutionary conservation and wide tissue distribution suggest an essential biological role. Its physical state, which is controversially discussed in previous publications, was investigated using small-angle X-ray scattering, dynamic light scattering, mass spectrometry, and circular dichroism (CD). Our results unequivocally demonstrate that prothymosin is a monomer under physiological conditions. The protein adopts a random coillike conformation but exhibits persistence of direction and curvature. No regular secondary structure is detectable by CD. The Stokes radius, Rs = 3.07 nm, and the radius of gyration, RG = 4.76 nm, are 1.77 and 3.42 times larger, respectively, than those expected for a compactly folded protein consisting of 109 amino acid residues. A remarkable amount of secondary structure is formed only in the presence of trifluoroethanol at low pH. The finding that a biologically active protein molecule with 109 amino acid residues adopts a random coil conformation under physiological conditions raises the question whether this is a rare or a hitherto-overlooked but widespread phenomenon in the field of macromolecular polypeptides.

Animals↗
PubMed1995

The thymus extract Thymex-L potentiates the retinoic acid-induced differentiation of the human myeloid leukemia cell line HL-60.

The human promyelocytic cell line HL-60 can be differentiated with retinoic acid (RA) along the granulocytic pathway. Numerous studies have identified many synergistic combinations of RA with cytostatics, cytokines and other inducers. A combination of RA with the crude thymus extract Thymex-L increased differentiation of HL-60 cells as confirmed by two functional assays and morphology, whereas the extract itself did not show any effect. The functional markers phagocytosis-associated chemiluminescence and nitroblue tetrazolium reduction were more enhanced (up to 4-fold with 1000 micrograms/ml Thymex-L) than morphology. The effect was found over a wide RA concentration range (10(-11) - 10(-6) M) and was dependent on extract concentration. The half-maximal induction of both functional markers was reached at 400 micrograms/ml. To achieve the same effect with the combination in comparison with RA alone, an RA dose reduction of about 100-fold was estimated. The effect was also seen when the cells were pretreated with the thymus extract for two days. The enhancement of RA action by Thymex-L was not correlated with an increase of extracellular or intracellular RA concentration. The active compound in Thymex-L is heat stable and bigger than 5 kDa as confirmed by gelfiltration. The defined thymus peptides thymosin alpha 1, prothymosin alpha 1 and thymopentin were unable to synergistically enhance HL-60 differentiation. These data suggest that the treatment with a thymus extract can increase the sensitivity of HL-60 cells for RA. This may have clinical implications.

Cell Differentiation↗
PubMed1995

Synergistic interactions between interferon beta and carboplatin on SK-MEL 28 human melanoma cell growth inhibition in vitro.

Carboplatin and interferon beta (IFN beta) were tested alone and in combination for their antiproliferative activity on the human melanoma cell line SK-MEL 28 in vitro. Cells were incubated for 4 days in the presence of carboplatin (0.1 mM and 0.1 microM) and interferon beta (5 pM and 5 nM) and cell growth inhibition was determined by the sulphorhodamin B assay. The antiproliferative effects of the drug combinations were analysed using Berenbaum's hyperplane theorem to determine additive, synergistic and antagonistic effects. IFN beta was found to be 10,000 times more active in inhibiting cell growth of SK-MEL 28 cells than carboplatin on the basis of IC50 values (IFN beta: IC50 = 1.24 nM, carboplatin: IC50 = 18.2 microM). The addition of IFN beta at 0.5 nM reduced the IC50 value of carboplatin 18.0-fold; with IFN beta at 0.05 nM a dose reduction of 1.84 was measured. At the carboplatin: IFN beta molar concentration ratios of 2000:1 and 6000:1, interaction indices (I) of 0.66 and 0.83 were determined respectively, indicating synergistic interactions between the two drugs. At higher carboplatin: IFN beta molar ratios (20,000:1 and 60,000:1) an additive interaction was observed (I = 1.07 and 1.20). However, further in vitro studies with several melanoma cell lines are necessary to evaluate the potential effectiveness of the drug combination of carboplatin and IFN beta for eventual clinical utilisation.

Carboplatin↗
PubMed1995

Isolation and partial characterization of basic proteinases from stem bromelain.

Crude bromelain extracts from pineapple stems (Ananas comosus) were fractionated by two-step FPLC-cation-exchange chromatography. At least eight basic proteolytically active components were detected. The two main components F4 and F5 together with the most active proteinase fraction F9 were characterized by SDS-PAGE, mass spectroscopy, multizonal cathodal electrophoresis, partial amino acid sequence, and monosaccharide composition analysis. F9 amounts to about 2% of the total protein and has a 15 times higher specific activity against the substrate L-pyroglutamyl-l-phenylanalyl-l-leucine-p-nitroanilide (PFLNA) than the main component F4. The molecular masses of F4, F5, and F9 were determined to 24,397, 24,472, and 23,427, respectively, by mass spectroscopy. Partial N-terminal amino acid sequence analysis (20 amino acids) revealed that F9 differs from the determined sequence of F4 and F5 by an exchange at position 10 (tyrosine-->serine) and position 20 (asparagine-->glycine). F4 and F5 contained fucose, N-acetylglucosamine, xylose, and mannose in ratio of 1.0:2.0:1.0:2.0, but only 50% of the proteins seem to be glycosylated, whereas F9 was found to be unglycosylated. Polyclonal antibodies (IgG) against F9 detected F4 and F5 with tenfold reduced reactivity. The pH optimum of F4 and F5 was between pH 4.0 and 4.5 and for F9 close to neutral pH. The kinetic parameters for PFLNA hydrolysis were similar for F4 (Km 2.30 mM, kcat 0.87 sec-1 and F5 (Km 2.42 mM, kcat 0.68 sec-1), and differed greatly from F9 (Km 0.40 mM, kcat 3.94 sec-1).

Amino Acid Sequence↗
PubMed1995

Prothymosin alpha 1 modulates lymphokine-activated killer cell activity and IL-2 production by peripheral blood lymphocytes from melanoma patients in vitro.

The effects of prothymosin alpha 1 (Pro alpha 1) on the natural killer (NK), lymphokine (IL-2)-activated killer (LAK) cell activity and the phytohaemagglutinin (PHA)-induced IL-2 secretion of peripheral blood T-lymphocytes (PBL) from 34 malignant melanoma patients of all clinical stages were studied in vitro. On average, melanoma patients showed lower NK and LAK cell activities than healthy donors. In particular, patients with metastases revealed an impaired NK cell activity. However, individuals showed a broad range of LAK cell sensitivity to Pro alpha 1 depending, among other factors, on the disease stage. LAK cell activities were not correlated to tumour stage. Patients' impaired LAK cell activity could be restored by Pro alpha 1. Only patients at stage II (regional metastases) responded to Pro alpha 1. The IL-2 secretion from PBL melanoma and healthy donors did not differ, Pro alpha 1 administration was without any significant effect. However, stage III (distant metastases) PBL expressed significantly lower IL-2 levels, compared to stage I (primary tumours). The highest IL-2 levels was found to be associated with tumour stage II. Pro alpha 1 enhanced the IL-2 secretion from stage I PBL. Therefore Pro alpha 1 administration abrogated the defective LAK cell activity and IL-2 secretion of PBL, mainly from patients at early melanoma stages.

Adult↗
PubMed1994

Bromelain protease F9 reduces the CD44 mediated adhesion of human peripheral blood lymphocytes to human umbilical vein endothelial cells.

The thiol protease bromelain has been shown to remove T-cell CD44 molecules from lymphocytes and to affect T-cell activation. We investigated the effect of a highly purified bromelain protease F9 (F9) on the adhesion of peripheral blood lymphocytes (PBL) to human umbilical vein endothelial cells (HUVEC). Preincubation of the lymphocytes with F9 reduced the adherence to about 20% of unstimulated and to about 30% of phorbol-dibutyrate (P(Bu)2) stimulated lymphocytes. Using flow cytometry, both crude bromelain and protease F9 reduced the expression of CD44, but not of LFA-1, on PBL. F9 was about 10 times more active than crude bromelain; at 2.5 micrograms/ml of F9 about 97% inhibition of CD44 expression was found. A mAb against CD44 was tested and found to block the F9-induced decrease in PBL-binding to HUVEC. The results indicate that F9 selectively decreases the CD44 mediated binding of PBL to HUVEC.

Antibodies, Monoclonal↗
PubMed1994

Thymosin alpha 1 effects, in vitro, on lymphokine-activated killer cells from patients with primary immunodeficiencies: preliminary results.

In patients with primary immunodeficiencies the role of natural killer (NK)- and lymphokine (IL-2)-activated killer (LAK)-cells is not yet satisfactorily established. Using a clonogenic assay with K562 leukemia target cells, we studied their NK- and LAK-cell activity in vitro. Moreover, the effect of thymosin alpha 1 (T alpha 1) on LAK-cell activity was studied in 11 patients with different immunodeficiencies. The results were compared with data of healthy controls (n = 11) and cord blood samples (n = 6). Common variable immunodeficiency patients demonstrated a mean LAK-cell activity of about 65% of normal controls and cord blood samples. The moderately reduced LAK-cell activity was not affected by T alpha 1. In the immunodeficient other patients, low levels of LAK-cell activity with a mean value of 10% of normal controls were seen. The mean LAK-cell activity could be improved by T alpha 1: three patients showed an improvement of their LAK-cell activity up to 25-30% after T alpha 1 administration in vitro, but in one case T alpha 1 was without any effect. Analysis of the expression of the surface markers CD8, CD16, CD57 and CD8/CD57 revealed that only CD16 positive lymphocytes were significantly less in immunodeficient patients. We found a linear correlation between LAK-cell activity and CD8/CD57 double positive lymphocytes in all patients. Our results demonstrate that suppressed LAK-cell activity from immunodeficient patients can be individually improved by T alpha 1. Further in vivo studies should evaluate thymic peptide immunotherapy for individual immunodeficient patients.

Adolescent↗
PubMed1994

The safety of vaginal prostaglandin F2 alpha for the stimulation of labour.

In an audit of 15,102 consecutive deliveries between 1986 and 1991, 3,168 labours were induced with intravenous oxytocin and 824 with 40 mg prostaglandin F2 alpha (PGF2 alpha) vaginal gel. Four hundred and twenty women received PGF2 alpha alone and 404 received PGF2 alpha followed by oxytocin. The main aim of the study was to audit the safety of PGF2 alpha gel to stimulate labour. There were no maternal or neonatal complications attributable to this therapy. In particular, there were no cases of uterine rupture or hyperstimulation requiring surgical or pharmacological intervention. There was little difference in the evidence for fetal distress between induction methods. Although the prostaglandin and oxytocin groups were not comparable in all respects, the results of this large retrospective study confirmed the results of smaller prospective randomized trials showing a significantly shorter labour and reduced analgesia, surgical delivery and postpartum haemorrhage rates in women treated with PGF2 alpha alone. This is the largest reported series of PGF2 alpha induced labours and provides evidence of its safety and is in keeping with physiological data suggesting that PGF2 alpha is the main prostaglandin and oxytocic associated with normal progressive labour. Its apparent safety and potential to reduce both intervention in labour and postpartum complications merits greater attention.

Adult↗
PubMed1993

Continuous low-dose oestrogen and progestogen hormone replacement therapy: a randomised trial.

OBJECTIVE: To establish the efficacy and acceptability of combined continuous low-dose oestrogen and low-dose progestogen therapy, to determine whether any of three commercially available progestogens had any advantages or disadvantages in these circumstances and whether use of the lowest clinically effective oestrogen dose affected other outcomes being measured. DESIGN: A 12-month, prospective, open label, single centre, randomised trial. PATIENTS AND METHODS: Seventy-five postmenopausal women already receiving hormone replacement therapy in the form of conjugated equine oestrogens (CEE) (0.625 mg daily) and cyclical medroxyprogesterone acetate (10 mg) and experiencing withdrawal bleeding were changed to a continuous daily regimen of 0.3 mg CEE and a random allocation of one of three low-dose progestogens (medroxyprogesterone acetate 2.5 mg, levonorgestrel 30 micrograms or norethisterone 350 micrograms). Return to a dose of 0.625 mg CEE was permitted if required to control menopausal symptoms with separate analysis of this group when appropriate. OUTCOMES MEASURED: Menopausal symptom score, clinical bleeding pattern, endometrial biopsy results, forearm bone density and content, serum lipids and side effects. RESULTS: Fifteen women withdrew from the trial, five because of irregular bleeding. In the remainder, amenorrhoea was achieved in 53% by three months, in 67% by six months and in 93% by 12 months. Endometrial biopsy showed atrophic endometrium by 12 months in all but one patient, in whom minimal proliferative activity was seen. Twenty-seven women chose to return to a dose of 0.625 mg CEE. In all groups, final control of menopausal symptoms improved. All regimens were bone sparing and the lipid profile was unchanged. Minimal side effects were experienced by the patients. There was little difference in outcome between the three progestogens except that norethisterone therapy was associated with a greater prevalence of amenorrhoea at six months than was seen in the levonorgestrel and medroxyprogesterone acetate groups. CONCLUSIONS: These low-dose continuous oestrogen and progestogen regimens appear an appropriate option for the postmenopausal woman wishing to eliminate withdrawal bleeding and reduce both hormonal side effects and menopausal symptoms. The long term benefits of these regimens with regard to the prevention of osteoporotic fractures, cardiovascular disease and endometrial cancer need to be further assessed over time.

Bone Density↗