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Biomedical subjects

H Lahm

Publications and source records attributed to H Lahm.

At least 37 records · Page 2Linked to original sources
PubMed2000

Long-term survival in small cell lung cancer patients is correlated with high interleukin-2 secretion at diagnosis.

PURPOSE: We have previously reported significant impairment of IL-2 secretion in patients with small cell lung cancer (SCLC) at the time of diagnosis. Impairment of IL-2 secretion correlated with reduced survival in SCLC. This new prognostic factor was independent of other factors of prognostic relevance in SCLC. The prognostic value of IL-2 secretion was comparable to the most predominant prognostic factors for survival in SCLC identified so far. We now report long-term survival data from these patients. METHODS: The significance of correlations between single parameters in the test groups was calculated by using linear regression analysis, the Wilcoxon rank sum test, and Fisher's exact test. Using the Kaplan-Meier method, the log-rank test and the Cox regression model, we analyzed the relation of IL-2 secretion in whole blood cell cultures from 52 patients with SCLC at the time of diagnosis to established prognostic factors relevant for survival in SCLC. RESULTS: IL-2 secretion correlates with survival in SCLC. In addition, survival analysis according to tumor response and level of IL-2 secretion at the time of diagnosis demonstrates that long-term survival can only be observed after complete response to chemotherapy and high initial IL-2 secretion. CONCLUSIONS: IL-2 secretion at the time of diagnosis represents an independent prognostic factor for survival in SCLC. Moreover, long-term survival is only observed in patients with complete response upon chemotherapy that showed high IL-2 secretion at diagnosis. Therefore, IL-2 secretion may partially define long-term survival in this disease. These results have to be confirmed in a larger patient population.

Adult↗
PubMed2000

Effects of IGFBP-2 overexpression in vitro and in vivo.

Insulin-like growth factor-binding proteins (IGFBPs) are important modulators of IGF actions and may have both stimulatory and inhibitory effects. Expression of IGFBP-2 is increased after fasting and in a variety of pathological conditions. However, the specific role of IGFBP-2 in growth physiology remains to be determined. In this review, we summarize data from in vitro and in vivo models suggesting that IGFBP-2 has mainly inhibitory effects on IGF actions. Since the growth hormone (GH)/IGF system is involved in a number of pathological alterations of the kidney and these changes may--at least in part--be due to increased IGF-I, local overexpression of inhibitory IGFBP-2 in the kidney might prevent IGF-I-induced lesions. This hypothesis will be tested by crossing GH transgenic mice, a common model of glomerulosclerosis, with transgenic mice characterized by systemic and renal overexpression of IGFBP-2.

Animals↗
PubMed2000

Transgenic mouse models for studying the functions of insulin-like growth factor-binding proteins.

The insulin-like growth factor-binding proteins (IGFBPs) comprise a family of six related peptides that interact with high affinity with IGFs. IGFBPs compete with IGF receptors for IGF binding, and as a consequence of this competition they can affect cell growth. In addition, IGF-independent regulatory mechanisms of IGFBPs have been described. Despite their common property to interact with IGFs every IGFBP is expressed in a tightly regulated time- and tissue-specific manner suggesting that each protein may have its own distinct functions. Several transgenic mouse models overexpressing IGFBP-1, -2, -3, or -4 were developed in the past few years. Brain abnormalities were a common feature of IGFBP-1 transgenic models. Individual strains showed alterations in glucose homeostasis, reproductive performance, and a reduction of somatic growth as the most prominent phenotypes. The latter was also the main effect observed in IGFBP-2 transgenic mice. The overexpression of IGFBP-3 under the control of an ubiquitous promoter resulted in selective organomegaly, whereas mammary gland-targeted expression of this protein caused an altered involution after pregnancy in this organ. Tissue-specific overexpression of IGFBP-4 resulted in hypoplasia and reduced weight of smooth muscle-rich tissues such as bladder, aorta, and stomach. This review summarizes the current knowledge about the actions of IGFBPs in vivo based on the presently established transgenic mice.

Animals↗
PubMed2000

Gene expression of galectin-9/ecalectin, a potent eosinophil chemoattractant, and/or the insertional isoform in human colorectal carcinoma cell lines and detection of frame-shift mutations for protein sequence truncations in the second functional lectin domain.

The family of Ca2+-independent galactoside-binding lectins with the beta-strand topology of the jelly-roll, referred to as galectins, is known to mediate and modulate a variety of cellular activities. Their functional versatility explains the current interest in monitoring their expression in cancer research, so far primarily focused on galectin-1 and -3. Tandem-repeat-type galectin-9 and its (most probably) allelic variant ecalectin, a potent eosinophil chemoattractant, are known to be human leukocyte products. We show by RT-PCR with primers specific for both that their mRNA is expressed in 17 of 21 human colorectal cancer lines. As also indicated by restriction analysis, in addition to the expected transcript of 571 bp an otherwise identical isoform coding for a 32-amino acid extension of the link peptide was detected. Positive cell lines differentially expressed either one (7 lines) or both transcripts (10 lines). Sequence analysis of RT-PCR products, performed in four cases, allowed to assign the standard transcript to ecalectin in the case of SW480 cells and detected two point mutations in the insert of the link peptide-coding sequence in WiDr and Colo205. Furthermore, this analysis identified the insertion of a single nucleotide into the coding sequence generating a frame-shift mutation, an event which has so far not been reported for any galectin. This alteration encountered in both transcripts of the WiDr line and the isoform transcript of Colo205 cells will most likely truncate the protein part within the second (C-terminal) carbohydrate recognition domain. Our results thus reveal the presence of mRNA for a galectin-9-isoform or a potent eosinophil chemoattractant (ecalectin) or a truncated version thereof with preserved N-terminal carbohydrate recognition domain in established human colon cancer cell lines.

Adenocarcinoma↗
PubMed1999

Genetic capsid modifications allow efficient re-targeting of adeno-associated virus type 2.

The human parvovirus adeno-associated virus type 2 (AAV2) has many features that make it attractive as a vector for gene therapy. However, the broad host range of AAV2 might represent a limitation for some applications in vivo, because recombinant AAV vector (rAAV)-mediated gene transfer would not be specific for the tissue of interest. This host range is determined by the binding of the AAV2 capsid to specific cellular receptors and/or co-receptors. The tropism of AAV2 might be changed by genetically introducing a ligand peptide into the viral capsid, thereby redirecting the binding of AAV2 to other cellular receptors. We generated six AAV2 capsid mutants by inserting a 14-amino-acid targeting peptide, L14, into six different putative loops of the AAV2 capsid protein identified by comparison with the known three-dimensional structure of canine parvovirus. All mutants were efficiently packaged. Three mutants expressed L14 on the capsid surface, and one efficiently infected wild-type AAV2-resistant cell lines that expressed the integrin receptor recognized by L14. The results demonstrate that the AAV2 capsid tolerates the insertion of a nonviral ligand sequence. This might open new perspectives for the design of targeted AAV2 vectors for human somatic gene therapy.

Amino Acid Sequence↗
PubMed1999

Overexpression of insulin-like growth factor-binding protein-2 in transgenic mice reduces postnatal body weight gain.

Insulin-like growth factor (IGF)-binding protein-2 (IGFBP-2) has been shown to inhibit IGF-dependent cell proliferation in a number of in vitro studies. However, no in vivo model of IGFBP-2 overexpression has been established so far. Therefore, we have generated transgenic mice, in which expression of a mouse IGFBP-2 complementary DNA is controlled by the cytomegalovirus (CMV) promoter. In two independent transgenic strains, transgene expression was highest in pancreas and stomach, followed by skeletal muscle, heart, colon, spleen, adipose tissue, brain, and kidney. Within the pancreas, IGFBP-2 expression was found in the islets but not in the exocrine part. Serum IGFBP-2 levels of CMV-IGFBP-2 transgenic mice were about 3-fold (P < 0.05) increased, compared with controls, whereas serum levels of IGF-I and IGF-II were unaffected by IGFBP-2 overexpression. Fasted serum glucose and fasted insulin levels were slightly reduced in transgenic mice, compared with controls. Postprandial serum glucose insulin levels were not affected by the genotype. At days later than 23, body weights of transgenic mice were significantly (P < 0.05) reduced in both sexes, compared with nontransgenic littermates. This reduction in body weight was mainly attributable to significantly (P < 0.05) lower carcass weights of CMV-IGFBP-2 transgenic vs. control mice. In contrast, absolute organ weights were not (or only as a tendency) reduced, except for the weight of the spleen, which was significantly (P < 0.05) lower in male transgenic than in control mice. Our data suggest that IGFBP-2 represents a negative regulator of postnatal growth in mice, potentially by reducing the bioavailability of IGF-I.

Animals↗
PubMed1999

Postnatal overexpression of insulin-like growth factor II in transgenic mice is associated with adrenocortical hyperplasia and enhanced steroidogenesis.

The influence of postnatal insulin-like growth factor II (IGF-II) overexpression on adrenal growth and function was investigated in 3-month-old male phosphoenolpyruvate carboxykinase (PEPCK) promoter human IGF-II transgenic mice, which are characterized by 4-to 6-fold elevated postnatal IGF-II serum levels. Plasma corticosterone levels of PEPCK-IGF-II transgenic mice were 2-fold higher than in age- and sex-matched controls, both in the morning (7.4 +/- 1.5 vs. 17.8 +/- 3.9 ng/ml, P < 0.01) and in the evening (33.3 +/- 6.5 vs. 65.3 +/- 12 ng/ml, P < 0.01). When PEPCK-IGF-II transgenic mice were subjected to an ACTH challenge, corticosterone levels were stimulated 6-fold, to 396 +/- 17 ng/ml after 60 min, compared with 230 +/- 24 ng/ml in the control group. In contrast to corticosterone, plasma ACTH levels were similar in transgenic and control mice, excluding an indirect effect of IGF-II at the hypothalamic or pituitary level. In vitro, the basal and ACTH-induced corticosterone production of adrenal glands from transgenic mice was higher (2-fold and 1.8-fold, respectively) than that of control organs. However, when normalized for adrenal weight, the in vitro corticosterone secretion was similar in both groups. At autopsy, adrenal weights of transgenic mice were significantly greater than those of control adrenal glands (3.3 +/- 0.2 vs. 2.0 +/- 0.2 mg, P < 0.01, n = 10). Furthermore, a local expression of human IGF-II could be demonstrated in transgenic adrenal glands by RT-PCR, whereas in normal adult mice, no adrenal expression of IGF-II was detected. Stereological investigation of adrenal glands from another set of PEPCK-IGF-II transgenic mice and controls (6-month-old males) demonstrated that the increase in adrenal weight in transgenic mice is mainly caused by a 50% increase in the number of zona fasciculata cells, whereas cell volume and zonation of transgenic adrenal glands remained unchanged. In conclusion, our data indicate that postnatal overexpression of IGF-II induces an increased adrenal weight and elevated corticosterone serum levels, presumably by a direct mitogenic effect of IGF-II on adrenocortical fasciculata cells.

Adrenal Glands↗
PubMed1998

Insulin-like growth factor-binding protein-2 inhibits proliferation of human embryonic kidney fibroblasts and of IGF-responsive colon carcinoma cell lines.

So far, the physiological role of insulin-like growth factor binding protein-2 (IGFBP-2) has not been demonstrated directly. Therefore, we transfected 293 cells with an expression vector containing the CMV promoter and the complete cDNA of mouse IGFBP-2. Secretion of bioactive IGFBP-2 into conditioned medium was demonstrated by Western ligand and Western immunoblotting and quantified by specific RIA. For the analysis of cell proliferation three clones exhibiting either high or low/no IGFBP-2 expression were selected and compared to non-transfected parental 293 cells. IGFBP-2 secreting clones displayed reduced conversion of thiazolyl blue when compared to negative clones or non-transfected parental 293 cells (P < 0.01). The lower growth activity measured in the IGFBP-2 secreting clones was compensated in great part by the administration of exogenous IGF-I or -II. Conditioned media of IGFBP-2 secreting clones inhibited growth of IGF-responsive colon tumor cell lines (LS513, HT-29) while those of negative clones did not. In addition, conditioned medium from a clone expressing high levels of IGFBP-2 inhibited anchorage-independent growth of LS513 and HT-29 cells. In contrast, growth of an IGF-unresponsive tumor cell line (Co-115) was not affected by the conditioned media. We hypothesize that IGFBP-2 might sequester the IGFs and thus prevent them from transferring their mitogenic signals.

Animals↗
PubMed1998

Identification of transgenic mice by direct PCR analysis of lysates of epithelial cells obtained from the inner surface of the rectum.

Conventional screening protocols for transgene integration in mice employ tail tips or blood samples as sources to obtain genomic DNA preparations. We have developed a simple alternative non-surgical method. Epithelial cells are scraped off the inner surface of the rectum with a sterile plastic inoculation loop and are lysed with Kawasaki buffer. The lysate can be directly examined in a polymerase chain reaction (PCR) analysis without any need for further DNA purification. This procedure causes minimal harm and stress to the animals and repeated samples can be obtained as often as necessary. This technique has been used successfully to identify transgenic mice from a number of different lines. The method allows quick screening of numerous animals and contributes to a reduction of the number of surgical biopsies required.

Animals↗
PubMed1998

Selective suppression of cytokine secretion in whole blood cell cultures of patients with colorectal cancer.

We have investigated the secretion of interferon alpha (IFN-alpha), IFN-gamma, interleukin-1alpha (IL-1alpha), IL-1beta, IL-2 and tumour necrosis factor alpha (TNF-alpha) in whole blood cell cultures (WBCCs) of colorectal cancer patients upon mitogen stimulation. Whereas the values for IL-1beta and TNF-alpha remained virtually unchanged in comparison with healthy control subjects, WBCCs of colorectal cancer patients secreted significantly lower amounts of IFN-alpha (P < 0.005), IFN-gamma (P < 0.0001), IL-1alpha (P < 0.0001) and IL-2 (P < 0.05). This reduction correlated with the progression of the disease. The total leucocyte and monocyte population were almost identical in both groups. In contrast, a dramatic depletion of lymphocytes was observed in colorectal cancer patients, which affected both lymphocyte counts (P < 0.0005) and their distribution (P < 0.0001). Our results suggest a selective suppression of cytokines in colorectal cancer patients that is related to tumour burden. Several mechanisms might account for this phenomenon, one of which might be lymphocyte depletion.

Adult↗
PubMed1998

Malignant transformation of simian virus 40-immortalized human milk epithelial cells by chemical carcinogenesis accompanied by loss of heterozygosity on chromosome 1 but not microsatellite instability.

Simian virus 40-immortalized human milk epithelial cells (HuMI) are anchorage dependent and non-tumorigenic but can spontaneously progress to anchorage-independent and tumorigenic cells. To see whether HuMI cells can be transformed into anchorage-independent cells by chemical carcinogens, we treated them with 3-methylcholanthrene (MCA, 10 microg/mL). After 7-8 wk of culture, none of the treated cells grew in soft agar. However, when HuMI cells treated with MCA were cultured with 12-O-tetradecanoylphorbol-13-acetate (TPA, 10 ng/mL), they grew in soft agar; cells treated with TPA alone did not. TPA at this dose was cytotoxic to HuMI cells but not to their tumorigenic subline HuMI-TTu2. The response of the anchorage-independent HuMI-T cells was intermediate. These results indicate that HuMI cells can be transformed by treatment with MCA plus TPA, possibly because TPA selects those cells that are progressing toward malignancy. All five clones from MCA plus TPA-induced transformed cells formed malignant carcinomas in nude mice. When microsatellite changes at 17 loci in HuMI, HuMI-T, HuMI-TTu2, and five MCA plus TPA-transformed cells were examined, none of these cell lines showed instability at any locus, and no change in microsatellite length was found. However, all five MCA plus TPA-transformed cell lines showed loss of heterozigosity at 1q21-23 and 1q42 loci. This region of chromosome 1 is known to contain at least one antiproliferative gene, and our results suggest that inactivation of such a gene may be essential for full transformation of HuMI cells by chemical carcinogens.

Breast↗
PubMed1997

Tumour-derived, endocrine, exogenous and therapeutic factors differentially modulate cytokine secretion in whole blood cell culture.

Following our previous results which showed that TGF-beta 1 suppressed the secretion of certain cytokines, we investigated the effects of different endogenous and exogenous factors on cytokine secretion in whole blood cell culture by using an enzyme-linked immunosorbent assay (ELISA) for measurement of cytokine concentrations. Several molecules including dexamethasone, noradrenaline (NA) and ethanol differentially inhibited mitogen-induced cytokine secretion. Dexamethasone and noradrenaline suppressed secretion of IL-2, IFN alpha, IFN gamma, TNF alpha, IL-1 alpha and IL-1 beta. beta-Endorphin and Leu-Enkephalin had no significant influence on cytokine secretion. Suppression of cytokine secretion by TGF-beta 1 was further intensified significantly and dose dependently by addition of noradrenaline. GM-CSF stimulated the secretion of IL-1 alpha, IL-1 beta and TNF gamma, but had no influence on the secretion of IL-2, IFN alpha and IFN gamma. G-CSF, IL-3 and SCF did not significantly influence secretion of all cytokines tested. Thus, endogenous and exogenous factors differentially influence cytokine secretion by immunocompetent cells.

Adult↗
PubMed1997

Decrease of interleukin-2 secretion is a new independent prognostic factor associated with poor survival in patients with small-cell lung cancer.

BACKGROUND: We have previously shown that suppression of interleukin-2 (IL-2) secretion was mediated by transforming growth factor (TGF) beta 1 secreted by small-cell lung cancer (SCLC) tumor cells. We have also shown that IL-2 secretion was significantly impaired in patients with SCLC at the time of diagnosis. Reconstitution of cytokine secretion correlated with reduction of tumor load. These data suggested that the immune system was suppressed by the tumor. To address the clinical relevance of cytokine suppression in SCLC, we investigated the correlation of the level of IL-2 secretion with survival. PATIENTS AND METHODS: The significance of correlations between single parameters in the test groups was calculated by using the linear regression analysis, the Wilcoxon rank sum test and the exact test according to Fisher. Using the Kaplan-Meier method, the log-rank test and the Cox-regression model, we analysed the relation of IL-2 secretion in whole blood cell cultures from 52 patients with SCLC at the time of diagnosis to established prognostic factors relevant for survival in SCLC. RESULTS: Impairment of IL-2 secretion significantly correlates to survival in SCLC (P = 0.004). Further univariate and multivariate analysis showed that this prognostic factor is independent from other factors of prognostic relevance in SCLC, namely stage of disease, neurone specific enolase (NSE), lactate dehydrogenase (LDH), age, and sex. More important, the prognostic value of IL-2 secretion is comparable to the most predominant prognostic factors for survival in SCLC identified so far. In the final model of the cox regression analysis, the P-value for IL-2 secretion in relation to stage of disease was 0.012 and 0.019, respectively. CONCLUSIONS: IL-2 secretion at the time of diagnosis represents an independent prognostic factor for survival in SCLC. Although its prognostic value has to be confirmed in a larger group of patients, our results demonstrate that IL-2 secretion may play an important role in diagnosis and treatment of SCLC. Moreover, in contrast to other prognostic factors, impairment of IL-2 secretion may help to understand immunosuppression in SCLC and, thus, important elements of the pathogenesis of this disease.

Adult↗
PubMed1997

Role of integrins and evidence for two distinct mechanisms mediating human colorectal carcinoma cell interaction with peritoneal mesothelial cells and extracellular matrix.

Peritoneal carcinomatosis involves a series of events including tumor cell interactions with mesothelial cells and the extracellular matrix (ECM). We have studied the adhesive and invasive properties of four human colorectal carcinoma cell lines (Co115, HT29, SW480, SW620) confronted in vitro with a human mesothelial cell monolayer or with the ECM proteins collagen IV, laminin-1, fibronectin, tenascin-C and vitronectin. Quantitation was achieved following staining of tumor cells with the calcein-AM fluorescent dye. We found that all four cell lines rapidly adhered to a mesothelial cell monolayer. This adhesion event was not inhibitable by anti-integrin and anti-CD44 antibodies. Following initial attachment, the SW480 and SW620 cells invaded the mesothelial cell monolayer more aggressively than HT29 and Co115 cells. All cell lines adhered to ECM proteins with each one exhibiting an individual adhesion pattern. Adhesion to matrix was completely integrin-dependent. When tested in an invasion assay, HT29 and Co115 cells crossed Matrigel-coated filters while SW480 and SW620 cells did not. This invasion was inhibited by anti-beta 1 integrin antibodies. Taken together, our results demonstrate that the initial colorectal tumor cell-mesothelial cell interaction occurs through an integrin-independent mechanism while adhesion to matrix proteins and invasion through Matrigel are integrin-dependent events. Furthermore, the different invasive capacity of SW480 and SW620 versus HT29 and Co115 cells upon interaction with a mesothelial cell monolayer or Matrigel suggests that these two invasion events may be mediated by distinct mechanisms.

Cell Adhesion↗
PubMed1996

Growth inhibition signalled through the interleukin-4/interleukin-13 receptor complex is associated with tyrosine phosphorylation of insulin receptor substrate-1.

Induction of growth inhibition in human colorectal carcinoma cell lines by interleukin (IL)-4 and IL-13 was associated with the neophosphorylation of a 170 kDa cellular protein, identified as insulin receptor substrate-1 (IRS-1) by immunoprecipitation. Tyrosine phosphorylation of IRS-I was also induced by insulin and insulin-like growth factor I. Sublines of colorectal carcinoma cells unresponsive to growth modulation by IL-4, IL-13 or insulin-like growth factor I-induced growth did not phosphorylate IRS-1. A functional, multimeric IL-4 receptor complex was present on all carcinoma cell lines with a subunit composition of 65 kDa, 75 kDa and the previously characterized 130 kDa band as demonstrated by affinity cross-link with 126I labelled IL-4. The 65 kDa subunit is novel whereas the 75 kDa band represents the common IL-2 receptor gama-chain the novel 65 kDa receptor was present as a double band and bound primarily 125I-labelled IL-13. The present study demonstrates the involvement of a novel chain other than the gama-chain in the receptor complexes of IL-4 and IL-13 and and post-receptor tyrosine phosphorylation of IRS-1. The association of IRS-1 with growth inhibitory signals in carcinoma cells suggests a novel mechanism of tumour growth control.

Antigens, CD↗
PubMed1996

Differential effect of interleukin-4 and transforming growth factor beta 1 on expression of proto-oncogenes and autocrine insulin-like growth factor II in colorectal carcinoma cells.

We have compared the effect of interleukin-4 and transforming growth factor beta 1 on proliferation and gene expression in two colorectal carcinoma cell lines, LS513 and LS1034. Transforming growth factor beta 1 was a potent inhibitor for both cells lines and virtually abolished de novo DNA synthesis. Interleukin-4 inhibited thymidine incorporation up to 60 and 45%, respectively. While both cytokines exerted a comparable cyto-inhibitory activity they displayed differential effects on proto-oncogene expression. Transforming growth factor beta 1 markedly down-regulated c-myc in LS1034 but not in LS513 cells. In contrast, expression of c-fos was induced by interleukin-4 in LS513 but not in LS1034 cells. Interestingly, in agreement with their cyto-inhibitory activity both cytokines suppressed the expression of insulin-like growth factor II in LS1034, which is an autocrine growth factor for these cells.

Cell Division↗