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H Kirchner

Publications and source records attributed to H Kirchner.

At least 217 records · Page 12Linked to original sources

The long persistence of CMV DNA in the blood of renal transplant patients after recovery from CMV infection.

A total of 30-50% of all renal transplant recipients undergo infections caused by human cytomegalovirus. With the introduction of ganciclovir and foscarnet for specific antiviral therapy there is an increasing demand for diagnostic tools that allow the early and rapid identification of CMV as the causative agent of the observed disease. We and others previously showed the direct detection of pp65 antigen in peripheral blood leukocytes to be an excellent marker for active cytomegalovirus infection. In order to establish whether the detection of CMV DNA by the polymerase chain reaction (PCR) supplies further information in this regard, we compared both methods. In 41 renal transplant patients the PCR assay yielded a sensitivity of 100% compared with 87.5% of the antigenemia assay. Specificities reached 67% and 92.5%, respectively. In 5 patients without both serological signs of infection and antigenemia, CMV DNA was also found. The duration of CMV DNA detection in PBL during active infection was significantly longer than antigenemia. Even after successful treatment of symptomatic CMV disease, DNA was present for a period of weeks without any relapse of disease. In contrast, antigenemia disappeared after antiviral therapy and reappeared only in one patient with relapse of CMV disease. We conclude that PCR offers no advantages over antigen detection in monitoring for CMV infections after renal transplantation.

Base Sequence↗

HLA DNA preorgan retrieval donor typing in renal transplantation.

For the first time, a DNA technique was used in clinical practice for pre-organ donor retrieval typing in renal transplantation. The method is based on the polymerase chain reaction with nested sequence-specific pairs (nested PCR-SSP) for all serological HLA-DR specificities of DR1-DR18. A panel of reference-typed individuals (n = 101) was investigated in a blind quality control study and the results revealed a sensitivity of 99.5% and a specificity of 100%. Semi-automation of PCR-SSP provided rapid, accurate and reliable typing results demonstrating that this DNA test is most suitable for replacing the error-prone HLA-DR serological technique. Twenty-seven consecutive cadaveric organ donors have been successfully typed so far. The results were available within less than 3 h after blood sampling and in all cases prior to the retrieval of organs. In conclusion, the introduction of this accurate DNA typing in HLA matching programs may significantly improve the graft survival rate in renal transplantation.

Blood Grouping and Crossmatching↗

Helium-neon laser irradiation induces effects on cytokine production at the protein and the mRNA level.

The construction of an in vitro model allowed an investigation of the basic functions of immunocompetent cells after laser irradiation. Among low-energy laser sources, the helium-neon (He-Ne) laser, with a wavelength of 632.8 nm, has often been found to produce photobiological effects including evidence of interference with immunological functions. Previous experiments revealed an influence of He-Ne laser irradiation on concentrations of interleukin-1 alpha (IL-1 alpha), tumor necrosis factor-alpha (TNF-alpha), interleukin-2 (IL-2), and interferon-gamma (IFN-gamma) in supernatants of cultures of human peripheral blood mononuclear cells (PBMC) with increased cytokine concentrations after irradiation of 18.9 J/cm2 and decreased concentrations after irradiation of 37.8 J/cm2. Now, the mechanisms involved were studied. Results showed that cytokine production of cells stimulated with phytohemagglutinin (PHA), concanavalin A (Con A), or bacterial lipopolysaccharide (LPS) was altered significantly after laser irradiation but not after stimulation with staphylococcus aureus enterotoxin B (SEB). In situ hybridization of IFN-gamma mRNA producing PBMC revealed that the number of positive cells was modulated similarly. The results were identical in cultures of enriched monocytes (M phi) or enriched T cells. Cells of the human monocytic cell line Mono Mac 6 were also influenced after LPS stimulation, whereas constitutively IL-2-producing Jurkat cells were not influenced by laser irradiation at any energy density. Analysis of the IL-2 receptor (IL-2R) and intercellular adhesion molecule-1 (ICAM-1) expression in PBMC showed partial down-regulation of both receptors at 37.8 J/cm2, but only after stimulation with PHA.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Adhesion Molecules↗

Low-dose interleukin-2 in combination with interferon-alpha effectively modulates biological response in vivo.

Phenotypic characterization of peripheral blood lymphocytes was performed in patients with advanced metastatic cancer receiving low-dose recombinant interleukin-2 (rIL-2) and recombinant interferon-alpha (rIFN-alpha) as subcutaneous home therapy. A total of 31 patients with progressive metastatic renal cell carcinoma, malignant melanoma, colorectal cancer, B-cell lymphoma, and Hodgkin's disease, were evaluated. Patients were treated with a combination of low-dose subcutaneous rIL-2 and rIFN-alpha, consisting of a 2-day rIL-2 pulse at 9.0 million IU/m2 twice daily, followed by 6 weeks of combined low-dose rIL-2 at 1.8 million IU/m2 twice daily, 5 days per week, and rIFN-alpha at 5.0 million U/m2 3 times per week. This treatment regimen resulted in an overall significant (p < 0.002) increase in peripheral blood lymphocyte subsets expressing CD3, CD8, CD16, CD25, and CD56. Expansion of peripheral blood natural killer (NK) cells was correlated to treatment response. Thus, treatment-related increase in CD56-positive lymphocytes was 1.8-fold higher in complete or partial responders when compared to progressive disease patients (p = 0.0). Increase in NK cells upon low-dose rIL-2 and rIFN-alpha was associated with a significant expansion (p = 0.0) of peripheral blood eosinophils (r = 0.71). Patient pretreatment using rIL-2, rIL-2 and rIFN-alpha, or chemotherapy abrogated the treatment-induced induction of NK cells and IL-2 receptor- (CD25) positive T lymphocytes, respectively. Peripheral blood NK cells were significantly decreased (p < 0.05) in patients developing neutralizing antibodies specific to rIL-2.

Antibodies↗

Hematotoxicity of interleukin-2 in man: clinical effects and comparison of various treatment regimens.

BACKGROUND: Immunotherapy using recombinant human interleukin-2 (rIL-2) produces objective responses in a proportion of advanced cancer patients. While early investigators employed intravenous (i.v.) treatment regimens, recent clinical trials applied therapy schedules via subcutaneous (s.c.) injection, mostly in combination with recombinant human interferon-alpha (rIFN-alpha). There were no significant differences in reported response rates between i.v. and s.c. treatment regimens. METHODS: We retrospectively evaluated 148 treatment cycles of s.c. immunotherapy administered to 107 outpatients. Adverse effects of s.c. cytokine therapy were described with regard to hematotoxicity, and compared to adverse effects reported upon high- or intermediate-dose i.v. rIL-2 therapy. Our study population consisted of 15 patients who received s.c. rIL-2 at doses of 4.8-14.4 million IU/m2/day on 5 days per week, 20 patients who were treated with rIFN-alpha 2b at 3.0-6.0 million U/m2/day thrice weekly, and 72 patients who were given s.c. rIFN-alpha 2b at 6.0 million U/m2/day thrice weekly plus s.c. rIL-2 at 14.4-18.0 million IU/m2/day on days 1 and 2, followed by 4.8 million IU/m2/day, 5 days per week. RESULTS: Subcutaneous immunotherapy as administered in this study was well tolerated in the outpatient setting. Thus, there were no treatment-related deaths, and no patient developed grade III or IV (WHO) toxicity. Subcutaneous rIL-2 as a single agent produced grade II anemia and granulocytopenia in 7% of patients, respectively. In combination with s.c. rIFN-alpha, s.c. rIL-2 yielded grade II anemia and granulocytopenia in 8 and 14% of patients, respectively. In comparison, rIFN-alpha as a single agent produced grade III anemia and grade I granulocytopenia in 5 and 20% of patients, respectively. Upon s.c. rIL-2/rIFN-alpha combination therapy, a mean hemoglobin nadir of 114.4 g/l (p < 0.0001 when compared to baseline) was noted, and the mean granulocyte nadir was 2.3/nl (p < 0.0005). These mild laboratory changes due to cytokine-induced hematotoxicity were in marked contrast to severe anemia, thrombocytopenia, and granulocytopenia reported upon i.v. rIL-2 therapy. CONCLUSIONS: Palliative s.c. rIL-2-based immunotherapy as used in this study abrogates profound hematotoxicity, previously seen upon rIL-2-based i.v. immunotherapy. Subcutaneous rIL-2/rIFN-alpha combination therapy can be given in the ambulatory setting with good practicability and excellent safety. This outpatient regimen is as effective in advanced renal cell cancer as the most aggressive i.v. rIL-2-based protocols reported.

Adult↗

Expansion of peripheral blood natural killer cells correlates with clinical outcome in cancer patients receiving recombinant subcutaneous interleukin-2 and interferon-alpha-2.

Natural killer (NK) cells are believed to contribute to the clinical efficacy of cancer immunotherapy using recombinant interleukin-2 (rIL-2) in humans. In previous trials of high-dose i.v. rIL-2, however, no correlation has been established between circulating NK cells and treatment response. Between January 1989 and October 1990, we treated a total of 47 outpatients with advanced tumors using low-dose s.c. rIL-2 and interferon-alpha-2 (rIFN-alpha). Therapy consisted of a 2-day rIL-2 pulse at 18 million IU/m2/day, followed by 6 weeks of rIL-2 (3.6 x 10(6)-4.8 x 10(6) IU/m2/day x 5 days/week) and rIFN-alpha (5 x 10(6)-6 x 10(6) U/m2 x 3/week). Before and after therapy, we phenotypically evaluated circulating lymphocytes and correlated them with clinical response. During 6-week therapy, peripheral blood lymphocytes bearing the CD56 (NK-cell-associated) surface antigen were increased significantly (p < or = 0.005) in treatment responders [complete response (CR) and partial response (PR), n = 10; 3.8-fold] and stable disease (SD) patients (n = 20; 2.1-fold), while patients with progressive disease (PD, n = 17) exhibited no significant expansion of circulating NK cells (p > 0.1). After one 6-week treatment cycle, CR/PR patients had significantly more peripheral NK cells, when compared with patients in SD (1.6-fold) and PD (1.9-fold) (p < 0.04). The overall number of circulating lymphocytes was also increased upon therapy (1.6-fold; p < or = 0.001), but remained independent of response (p > 0.4). These data demonstrate that s.c. rIL-2 and s.c. rIFN-alpha produce a significant increase in peripheral blood NK cells; this expansion correlates significantly with treatment response in advanced tumor patients receiving long-term combination immunotherapy at outpatient doses.

Antigens, CD↗

Prognostic significance of Ki-67 immunostaining in nonmetastatic renal cell carcinoma.

PURPOSE: Fresh tissue samples from nonmetastatic renal cell carcinoma (RCC) patients were analyzed by Ki-67 immunostaining to determine the prognostic significance of this tumor-biologic parameter. MATERIALS AND METHODS: In a prospective study, Ki-67 immunostaining was performed on frozen sections of histologically proven node-negative RCC from 58 patients operated on between 1986 and 1988 to examine the method's prognostic value and its association with other clinicopathologic parameters such as tumor stage (pT) and grade (G). RESULTS: The percentage of Ki-67-positive cells (ie, the proliferation rate [PR]) of all 58 RCC tumors ranged between 1% and 23%, while normal renal tissue exhibited PRs up to 2% only. In almost all cases, the highest PRs were observed in the peripheral zone of malignant tissue close to the normal renal tissue. PR did not correlate with pT, whereas a strongly significant correlation was observed between PR and G, as well as recurrence rate. Twenty-three of 58 patients (39.6%) developed tumor recurrence. Disease-free survival was strongly associated with PR. In a multivariate analysis, G and PR were independent prognostic markers. CONCLUSION: The tumor-specific PR obtained by Ki-67 labeling seems to be an independent marker to describe the proliferative activity and aggressiveness of individual tumors. This new tumor-biologic marker detects RCC patients at high risk for recurrent disease, especially in those cases with identical pT and G.

Adult↗

T cells and their role in allergies.

The molecular and cellular mechanisms which up-regulate IgE synthesis in most atopic patients are only incompletely understood. There is no doubt that T cells play a prominent role in the B cell immunoglobulin switch to IgE. T-cell-derived interleukin (IL)-4 is a specific inducer of IgE synthesis which activates B cells to proliferate and, furthermore, induces other cells to release cytokines which enhance IgE synthesis. Additional costimulatory signals are required, however, for B cell activation and the consequent immunoglobulin switch to IgE. Besides physical B/T cell interaction, direct B cell activation through anti-CD40 provides such a signal. IL-4-induced B cell activation is amplified by other cytokines such as IL-5, IL-6, IL-3, probably IL-9, and tumor necrosis factor-alpha, whereas interferon (IFN)-gamma, IFN-alpha, transforming growth factor-beta and IL-12 are able to inhibit IL-4-induced IgE synthesis. These different IgE-synthesis-modulating lymphokines are secreted by different T cell subsets, TH1 and TH2. While IgE synthesis is activated by TH2-derived lymphokines, TH1 cell clones exhibit cytolytic activity. The detection of allergen-specific T cells in peripheral blood mononuclear cells (PBMNCs) of atopic patients, with a lymphokine profile similar to TH2, and the predominance of TH2 cells in PBMNCs of these patients might be one explanation for increased serum IgE concentrations in atopic individuals. It is not known, however, which stimulus leads to a predominant expression of TH2 cells over TH1 cells in atopic patients with consequently increased IgE synthesis.

B-Lymphocytes↗

[Interferon alpha antibodies show no cross reactions with typical autoantibodies].

Patients treated with natural human interferon alpha develop anti-interferon antibodies (IFN-AB) only in very rare cases. By contrast, patients with autoimmune disorders are able to generate high-titered IFN-AB against endogenous interferon alpha. One explanation for the development of auto-IFN-AB could be cross-reactivity with typical autoimmune antigens. We investigated the cross-reactivity of 3 high-titered IgG IFN-AB of female autoimmune patients (aged 32, 36, 74 years; two severe cases of SLE, one case of autoimmune thyroiditis) as well as 25 low-titered natural IgM IFN-AB of healthy blood donors (aged 19-48 years). Typical autoimmune antigens including dsDNA, ENA, as well as natural interferon beta and recombinant interferon gamma are not able to inhibit binding of IFN-AB to interferon alpha in an ELISA test system. Preincubation of sera containing either dsDNA antibodies (dsDNA-AB) (24 patients), thyroid peroxidase (TPO-AB) (9 patients) or thyroglobulin (TG-AB) (12 patients) with interferon alpha resulted in no change in the respective autoantibody titer. These data suggest that there is no cross-reactivity between IFN-alpha-AB and dsDNA-AB, TPO-AB or TG-AB. Thus, an explanation for the occurrence of IFN-AB in autoimmune disorders cannot be found in a cross-reaction between interferon alpha with typical autoimmune antigens.

Adult↗

Deficient interferon-alpha response of newborns in comparison to adults.

Human peripheral blood mononuclear cells (PBMC) respond efficiently to viral infections with interferon (IFN)-alpha production. Using Newcastle disease virus (NDV) and Sendai virus as inducers, our experiments with purified mononuclear cell populations of healthy adult volunteers showed that monocytes were the main IFN-alpha producers in these systems. Using an ELISA, which specifically recognized IFN-alpha 2, and a bioassay, in which all subtypes were detected, IFN-alpha 2 was a major subtype expressed. Furthermore, IFN-alpha release was higher after induction with NDV than after stimulation with Sendai virus. To compare the IFN-alpha response of adult PBMC to human cord blood mononuclear cells the cells were stimulated with NDV or Sendai virus. The IFN-alpha response of cord blood mononuclear cells to Sendai virus was slightly reduced compared to that of adult cells. However, IFN-alpha 2 was a major subtype produced. In contrast, when cord blood mononuclear cells were stimulated with NDV the IFN-alpha release was strongly diminished. Furthermore, IFN-alpha 2 was not the major subtype expressed. By in situ hybridization the number of IFN-alpha-producing cells was quantified. We found that 1% of adult PBMC and newborn leukocytes showed detectable IFN-alpha mRNA after stimulation with either Sendai virus or NDV. In conclusion, these data suggest that the ability of cord blood mononuclear cells to produce IFN-alpha is diminished compared to adult PBMC. This impaired IFN-alpha response is not based on a reduced number of IFN-alpha-producing cells, but rather due to underlying control mechanisms of the newborns that are different from the regulation of adult PBMC.

Adult↗

[Diagnosis of cytomegalovirus infections in immunosuppressed patients: the role of PCR].

The well-calculated and early application of specific antiviral drugs in cytomegalovirus (CMV) infections depends on sufficient virological diagnosis. Many authors prefer the detection of CMV DNA by the polymerase chain reaction (PCR) for this purpose. In our study in 41 renal transplant patients PCR yielded no additional information of clinical relevance compared to the already established pp65 antigenemia assay. However, neither of the two techniques allowed the prediction of symptomatic infections as also asymptomatic infections yielded positive results. In contrast, CMV mRNA was demonstrated by PCR mainly in symptomatic infections. We conclude, that mRNA amplification possibly is able to predict symptomatic CMV infections and will thereby allow the well-calculated application of e.g. ganciclovir.

Cytomegalovirus Infections↗

In situ hybridization of the mRNA for interferon-gamma, interferon-alpha E, interferon-beta, interleukin-1 beta and interleukin-6 and characterization of infiltrating cells in thyroid tissues.

Cytokine mRNA production in the thyroid tissues of patients with various thyroid diseases was analysed by in situ hybridization. In addition, infiltrating leukocytes were characterized by immunohistologic studies using the alkaline phosphatase anti-alkaline phosphatase (APAAP) staining technique. The following clinical material was investigated: two cases of Graves' disease, one with high and the other with a low amount of infiltrating leukocytes as well as two cases of non-toxic goitre also showing considerable quantities of infiltrating cells. The hybridization was performed on tissue sections with antisense probes for interferon-gamma (IFN-gamma), IFN-alpha E, IFN-beta, interleukin-6 (IL-6) and IL-1 beta. A small number of individual cells were found to express high levels of mRNA for IFN-gamma, IL-1 beta and measurable amounts of IL-6 throughout the tissue sections. However, IFN-alpha E or IFN-beta were not detected. Cytokine expressing cells were noted in the tissue of one patient with Graves' disease and in two cases with non-toxic goitre. In these samples a high amount of infiltrating leukocytes (CD45+) was detected, especially CD3+, CD8+, CD4+ and CD45RA+ T cells, in addition to B cells and macrophages. In one case an unusually large amount of T cell receptor gamma/delta+ (TcR gamma/delta+) cells was found. However, one sample of thyroid tissue derived from a patient with Graves' disease was poorly infiltrated and showed few cells expressing cytokines. In conclusion, using thyroid tissue as an example, our data suggest that the application of in situ hybridization with antisense RNA permits the study of cytokine production in tissues of both autoimmune and non-autoimmune origin.

Adult↗

Effects of cytokines on growth in vitro of primary human renal cell carcinoma.

In clinical trials different haematopoietic active cytokines such as granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) have been proven to alleviate myelosuppressive side effects of intensive chemotherapy in different non-urological malignancies. On the other hand, these cytokines can directly stimulate the proliferation of cells originating from some non-urological tumours. To clarify the impact of these cytokines on the proliferative behaviour of human renal cell carcinoma (RCC), 29 previously untreated RCC tumours were prepared for culturing in vitro using the cell cluster technique. The success rate for growth in vitro was 82.8% (24/29). The malignant renal cells were treated with different cytokines (GM-CSF, G-CSF and interleukin-3) in different dosages. Cell number and proliferation rates detected by immunostaining were used for treatment evaluation. A dosage-dependent stimulation of cell growth could not be observed compared to untreated cells. From the data presented in this study, proliferative stimulation of RCC by administering colony-stimulating factors in clinical trials cannot be assumed.

Carcinoma, Renal Cell↗

Cytokine production after helium-neon laser irradiation in cultures of human peripheral blood mononuclear cells.

The effects of laser light on the immune system have not been extensively characterized. Low-power laser sources, such as the helium-neon (He-Ne) laser with a wavelength of 632.8 nm, have been found to produce photobiological effects with evidence of interference with immunological functions. We have investigated the effects of He-Ne laser irradiation on Ficoll-Hypaque-isolated human peripheral blood mononuclear cells (PBMC). Cultured cells were irradiated for various times at two selected intensities and then stimulated with different mitogens. The rate of incorporation of 3H-thymidine into the DNA of stimulated cells decreased with increasing energy density. The levels of interleukin-1 alpha (IL-1 alpha), interleukin-2 (IL-2), tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) in supernatants of the cultures were determined (irradiated either before or after stimulation). When stimulating cells after irradiation, significantly increased levels of all cytokines were detected after 30 min of irradiation (18.9 J cm-2), whereas after 60 min of irradiation (37.8 J cm-2) cytokine levels were found to be significantly decreased.

Cell Division↗

Induction of cytokines in human whole blood cultures by a mitogen derived from Mycoplasma arthritidis and by staphylococcal enterotoxin B.

Mycoplasma arthritidis produces a so-far only partially characterized soluble material (MAS) that has a potent mitogenic effect on T lymphocytes of several species. Similar to staphylococcal enterotoxins and a number of related toxins secreted by other species of bacteria, nanogram quantities of these so-called superantigens are sufficient to induce significant amounts of cytokines in the supernatant of lymphocyte cultures. Induction of interleukin-6 (IL-6) by MAS in murine bone marrow-derived macrophages has recently been described. In our study, we examined the differential effects of MAS and Staphylococcus aureus enterotoxin B (SEB) on human blood cells. When compared to MAS, SEB induced a higher proliferative response and, accordingly, a higher release of IFN-gamma. In contrast, large amounts of the macrophage products IL-1, IL-6 and tumor necrosis factor alpha (TNF-alpha) were observed in supernatants of cell cultures stimulated with MAS, whereas only small amounts were induced by SEB. Staphylococci and mycoplasmas are responsible for a number of diseases with various symptoms in man and animals. Our results suggest that SEB and MAS show different qualities in lymphocyte and macrophage stimulation which may be relevant in the pathogenesis of diseases.

Antigens↗

Failure to detect human cytomegalovirus DNA in peripheral blood leukocytes of healthy blood donors by the polymerase chain reaction.

The transmission of cytomegalovirus (CMV) by blood transfusion may have a major effect on certain immunocompromised patients. To protect susceptible blood recipients from infection, it is advisable to use blood components from CMV-seronegative donors. However, serologic tests are not capable of indicating which blood component actually harbors infectious virus and can transfer it to the recipient. Therefore, a sensitive method is needed for the detection of the virus itself. There have been three reports on the detection of CMV in healthy volunteer blood donors by the polymerase chain reaction (PCR). CMV DNA was found in all seropositive and most seronegative blood donors. However, many other authors have failed to confirm these data. A highly sensitive and specific PCR assay was developed for the detection of CMV DNA in peripheral blood leukocytes. With this protocol, blood samples from 116 volunteer blood donors were investigated. None of these samples proved to be positive for CMV DNA. In contrast, CMV DNA was detected in 10 of 10 renal transplant patients early in the course of active CMV infection. It can be concluded that the CMV genome copy number in the peripheral blood leukocytes of healthy individuals is beyond the detection limit of current PCR technology.

Base Sequence↗