Biomedical subjects

S Linder

Publications and source records attributed to S Linder.

At least 73 records · Page 4Linked to original sources

PubMed1997

Signal transduction in fibroblasts stably transformed by [Val12]Ras--the activities of extracellular-signal-regulated kinase and Jun N-terminal kinase are only moderately increased, and the activity of the delta-inhibitor of c-Jun is not alleviated.

Ras-transformed cells often show high levels of expression of activating protein-1 and Ets and of genes regulated by these transcription factors. In analogy with the effects of transient stimulation of Ras, it is assumed that the increase in transcription-factor transactivation in stably transformed cells is due to Ras-induced constitutive activation of mitogen-activated protein kinases. However, this has not been extensively studied. Using specific substrate peptides, we have examined here the activities of two types of mitogen-activated protein kinase, extracellular-signal-regulated kinase (ERK) and Jun N-terminal kinase (JNK), in [Val12]Ras-transformed rat embryo fibroblast cell lines. These activities were elevated 2-3-fold in Ras-transformed cells compared with non-transformed cells with a similar growth rate. Increased ERK activity was not necessarily accompanied by a similar increase in JNK activity. In transformed cells, ERK and JNK activities could be stimulated fourfold and ninefold by phorbol ester and ultraviolet-light treatment, respectively, indicating that only a fraction of these enzymes were constitutively activated in these cells. It has been suggested that inactive JNK downregulates c-Jun transcriptional activity by binding to the c-Jun delta-domain. No decrease in delta-inhibitor activity could be demonstrated in Ras-transformed cells compared with control cells, consistent with the presence of mainly inactive JNK in transformed cells. Treatment of transformed cells wih benzodiazepine 5B, an inhibitor of Ras farnesylation, decreased ERK and JNK activities, and concomitantly caused morphological reversion, reduced growth rate, and normalization of transformation-related gene expression. We conclude that in stably Ras-transformed cells the moderately increased ERK/JNK activities are not coregulated, and that ERK rather than JNK activity correlated with transformation-related gene expression.

Animals↗

PubMed1997

Screening of germline mutations in the CDKN2A and CDKN2B genes in Swedish families with hereditary cutaneous melanoma.

BACKGROUND: Approximately 10% of human cutaneous melanomas occur in families in which several members are affected. The familial predisposition to this disease is often associated with dysplastic nevus syndrome, a condition in which afflicted family members have multiple dysplastic nevi (atypical moles). The chromosome region 9p21 and markers on chromosomes 1p and 6p have been linked to melanoma susceptibility. The tumor suppressor genes CDKN2A and CDKN2B have been mapped to the 9p21 region, and genetic analyses have revealed the presence of germline CDKN2A alterations in melanoma families. The reported frequencies of such alterations, however, vary among these families. PURPOSE: The present investigation was carried out to determine the frequencies of CDKN2A and CDKN2B germline gene mutations among members in a population-based cohort of Swedish melanoma families (i.e., melanoma kindreds). METHODS: DNA was prepared from blood samples obtained from 181 individuals belonging to 100 melanoma kindreds. The polymerase chain reaction (PCR) technique, followed by single-strand conformation polymorphism (SSCP) and nucleotide sequence analyses, were used to identify the types and frequencies of mutations in exons 1, 1beta, 2, and 3 of the CDKN2A gene and in exons 1 and 2 of the CDKN2B gene. RESULTS: CDKN2A gene aberrations were independently identified by both SSCP and nucleotide-sequence analyses. Nucleotide-sequence analysis identified a single point mutation leading to a substitution of leucine for proline in codon 48 of exon 1 in a family with a history of melanoma and several other cancers. A second abnormality, leading to an insertion of an extra arginine residue at codon number 113 of exon 2, was seen in four separate families. The CDKN2A exon-3 coding region had the wild-type sequence in all samples. No germline mutations were found in the alternative exon 1beta of the CDKN2A gene or in exons 1 and 2 of the CDKN2B gene. CONCLUSIONS: The present investigation demonstrates that CDKN2A germline gene mutations were observed in 7.8% of the 64 Swedish melanoma kindreds that each included at least two first-degree relatives with melanoma and dysplastic nevus syndrome. No CDKN2A exon 1beta or CDKN2B mutations were identified. The critical genes responsible for the inheritance of a susceptibility to develop melanoma among family members in this population have yet to be identified.

Carrier Proteins↗

PubMed1997

Sequence analysis and immunofluorescence study of alpha- and beta-tubulins in Reticulomyxa filosa: implications of the high degree of beta2-tubulin divergence.

We have cloned and sequenced 2 alpha- and 2 beta-tubulin isoforms from the giant freshwater amoeba Reticulomyxa filosa. The microtubules of this organism exhibit some unusual properties, including the highest rates of assembly and disassembly known and the inability to be stabilized by taxol. The cloned alpha-tubulins show a high degree of identity when compared to an alpha-tubulin consensus sequence. The beta-tubulins, however, are more divergent, the beta2-tubulin being the most unusual beta-tubulin found so far. The deduced amino acid sequence of beta2 shows 55% identity to a beta-tubulin consensus sequence. It also features 51 unique exchanges which cluster in the C-terminal half of the molecule. Several unique exchanges and two insertions occur in regions adjacent to, or directly implicated in, conserved beta-tubulin functions. A phylogenetic analysis places the beta-tubulins of R. filosa in the vicinity of beta-tubulins from fungi and slime molds. Monoclonal and polyclonal antibodies raised against R. filosa tubulins show that the electrophoretic mobility of alpha- and beta-tubulins is reversed with respect to tubulins from most other sources. Immunofluorescence experiments reveal a ubiquitous distribution of both beta-tubulins in the amoebal network. Our observations suggest possible links between the aberrant primary structure of the beta2-tubulin and the unusual properties of R. filosa microtubules.

Amino Acid Sequence↗

PubMed1997

Aspects of percutaneous fine-needle aspiration biopsy in the diagnosis of pancreatic carcinoma.

BACKGROUND: Morphologic verification of malignancy is usually a prerequisite before initiating nonsurgical therapy for pancreatic carcinoma. Fine-needle biopsy has been widely used, but the method has also been criticized because of inadequate sampling material and the risk of intra-abdominal seeding. METHODS: Cytologic material from 334 patients with suspected pancreatic malignancy was evaluated with regard to the sensitivity, specificity, and overall accuracy of the technique. The outcome (positive versus negative yield of cancer cells) was also entered in a Cox bivariate regression analysis and tested against multiple prognostic variables. RESULTS: In all, 270 patients had malignant disease, which was verified cytologically in 187. The sensitivity, specificity, and accuracy were 69%, 100%, and 75%, respectively. Statistically significant prognostic factors with regard to patient survival time were type of therapy (P <0.001), tumor location (P <0.001), jaundice (P <0.001), pain (P <0.001), and palpable mass (P <0.05). CONCLUSION: Fine-needle aspiration biopsy is still found to be a safe and feasible method for diagnosing pancreatic cancer, providing the requisite diagnostic information before initiating nonsurgical therapy.

Adult↗

PubMed1997

Distribution of stromelysin-3 mRNA transcripts and microvessels in human breast carcinomas.

Several proteinases, implicated in tumor invasion, are expressed in fibroblastic cells surrounding neoplastic cells, and are believed to be induced by paracrine stimulation. Such stimulation, by the local release of angiogenic factors, is also responsible for the induction of new capillary blood vessels, a crucial aspect of tumor growth and metastasis. We have here compared the expression of a matrix metalloproteinase, stromelysin-3 (ST3), with the distribution of tumor microvessels in invasive breast carcinomas. The highest level of ST3 mRNA in each tumor was recorded and compared to the highest microvessel density. No correlation between these two parameters was observed by analysis of 63 tumors. Detailed examination of 19 individual tumors did not reveal any correlation between the distribution of ST3 mRNA and microvessels. In the material studied here, ST3 expression was observed to correlate with long-term survival of the patients, whereas microvessel density did not correlate.

Breast Neoplasms↗

PubMed1997

Prognostic significance of Ki-67 antigen and p53 protein expression in pancreatic duct carcinoma: a study of the monoclonal antibodies MIB-1 and DO-7 in formalin-fixed paraffin-embedded tumour material.

Formalin-fixed paraffin-embedded material from 57 patients in whom curative resection of pancreatic carcinoma had been attempted was analysed by an immunohistochemical procedure to estimate proliferation and p53 protein expression. Using the monoclonal antibody (MAb) MIB-1, which recognizes a Ki-67 epitope, the proliferating cell index (PCI, percentage of immunoreactive tumour nuclei) and proliferating cell area (PCA, percentage of immunoreactive tumour nuclear area) were calculated using an interactive image analysis system and were compared with semiquantitative scoring of stainability. MAb DO-7, which recognizes both wild- and mutant-type p53 protein, was used to assess p53 expression in the same material. MIB-1 stainings were of high quality in 53 tumours. The median PCI was 29.7% (range 0.5-82.1%) and the median PCA was 10.6% (range 0.0-36.5%). There was a close correlation between PCI and PCA (P < 0.0001). PCI and PCA values were in conformity with the semiquantitative scoring (P < 0.0001). The p53 immunohistochemical stainings were successful in 48 tumours and the protein was expressed in 22 (46%). High PCI values (> 45%, n = 14) correlated with shorter survival time (P < 0.01). PCA (P < 0.05) and the expression of p53 protein (P < 0.001) were independent prognostic variables.

Aged↗

PubMed1997

Multicenter prospective evaluation of proposed pediatric migraine revisions to the IHS criteria. Pediatric Headache Committee of the American Association for the Study of Headache.

Eighty-eight children and adolescents were prospectively evaluated at eight specialty clinics comparing the diagnostic criteria of the International Headache Society (IHS) and the proposed revised (IHS-R) classification to the clinical diagnosis. The proposed revisions to the IHS classification for pediatric migraine include: duration-1 hour to 48 hours; location--bifrontal/bitemporal or unilateral; and symptoms--to include photophobia or phonophobia. A comparison of the diagnostic rates of pediatric migraine with and without aura of the total sample revealed IHS (66%) versus IHS-R (93%) and comparison of a subset of those patients less than 12 years of age (n = 39) revealed IHS (49%) versus IHS-R (87%). Significant improvement in the diagnostic sensitivity of migraine in the pediatric population was obtained by specific modifications to the IHS criteria pertaining to duration, location, and the symptoms of photophobia and phonophobia in an ongoing multicenter prospective study. These revisions may help to form the basis for future research guidelines and for further modifications to improve the diagnostic sensitivity of pediatric migraine maintaining the IHS model.

Adolescent↗

PubMed1997

Treatment of thromboembolic complications of fulminant meningococcal septic shock.

A patient report of fulminant meningococcal septic shock is described. The presentation, hospital course, and reconstructive efforts are outlined, and a brief review of meningococcal infection is included. Emphasis is placed on the algorithm used to determine treatment. A 19-year-old Hispanic male presented with all the hallmarks of Waterhouse-Friderichsen syndrome (WFS)-sudden onset, high fever, dyspnea with intermittent cyanosis, shock, disseminated intravascular coagulopathy, and the development of purpura. The pathognomonic feature of WFS-hemorrhage into the adrenal glands-if present, was not extensive, as he did not require steroid supplementation. Though cerebrospinal fluid latex agglutination was negative, his serum was positive for group C Neisseria and admission blood cultures grew Neisseria meningitidis. Thromboembolic complications were systemic with the highest morbidity peripherally in the lower extremities. Care for these injuries involved every rung of the reconstructive ladder-from local wound care and skin grafts to local flaps and microvascular transplantation.

Adult↗

PubMed1997

Inhibition of the growth of 12V-ras-transformed rat fibroblasts by acetylsalicylic acid correlates with inhibition of NF-kappa B.

Epidemiological studies have demonstrated a correlation between regular aspirin (acetylsalicylic acid; ASA) use and a decreased risk for the development of cancer. We here show that ASA inhibits the growth of 12V-ras-transformed rat fibroblasts in vitro at pharmacological concentrations. This effect appeared to be unrelated to inhibition of cyclooxygenase, since other cyclooxygenase inhibitors did not inhibit cell growth. A number of nuclear transcription factors have been implicated as mediators of transformation. ASA has recently been reported to inhibit the activation of one such factor, NF-kappa B. We found that NF-kappa B binding activity was decreased in ASA-treated 12V-ras-transformed cells. Inhibition of NF-kappa B activation was not due to a general inhibitory effect, since AP-1 binding activity was not affected. We conclude that ASA inhibits the growth of 12V-ras-transformed fibroblasts, possibly via inhibition of NF-kappa B.

Animals↗

PubMed1997

Surgical treatment of focal epilepsy in children: results in 37 patients.

This report concerns 37 children and teenagers operated upon for intractable seizures between 1990 and 1994. Follow-up is at least 3 years. Fourteen children underwent pure temporal lobe resections; 71% are seizure free, and 93% have a better than 90% decrease in seizure frequency. The presence of a lesion on magnetic resonance imaging, the side of the lesion, or the presence of abnormal pathology had no influence on the result of resection. 28% of the children who had extratemporal resections are seizure free, and 83% have a greater than 90% decrease in seizure frequency. There was a trend to better results in those with a lesion on magnetic resonance imaging. In the small group with temporal plus extratemporal foci, the results were poor with only 60% showing a greater than 90% reduction in seizure frequency.

Adolescent↗

PubMed1997

Codeletion of the JUN proto-oncogene and the CDKN2A tumor-suppressor gene in HRAS-transformed rat embryo fibroblast cell lines.

The cyclin kinase inhibitor p16, encoded by the CDKN2A gene, suppresses the transformation of mouse embryonic fibroblasts by oncogenic RAS. In contrast, the c-JUN transcription factor (a major component of AP-1) has been suggested to be required for RAS transformation of rodent fibroblasts. The CDKN2A gene and the JUN proto-oncogene have both been mapped to rat chromosome band 5q31-33. We here show that both copies of the CDKN2A gene are deleted in four of eight transformed cell lines derived from the transfection of rat embryo fibroblasts (REF) with HRASVAL12. In two cell lines, the homozygous deletions involved a larger area on 5q31-33, which included the JUN proto-oncogene. JUN-defective cells showed high AP-1 binding activity. Both AP-1 binding activity and stromelysin (transin) mRNA expression were found to be RAS-dependent in one of the JUN-defective cell lines. The finding of deletions of the CDKN2A gene in RAS-transformed REF cell lines is consistent with the concept that CDKN2A suppresses transformation by RAS. The occasional concomitant loss of the adjacent JUN proto-oncogene does not prevent establishment of transformed and tumorigenic cell lines.

Animals↗

PubMed1996

Assessment of homogeneity in polypeptide expression in breast carcinomas shows widely variable expression in highly malignant tumors.

We describe the results from a protein-based approach to the study of heterogeneity in gene expression between human tumors. Cell preparations from 5 benign breast lesions, 5 potentially weakly malignant and 4 potentially highly malignant invasive ductal breast carcinomas were examined by 2-dimensional gel electrophoresis (2-DE) gels. Qualitative and quantitative differences were recorded by computerized analysis. Analysis of samples from different areas of the same tumor showed a high degree of similarity in the pattern of polypeptide expression. Analysis of 2 tumors and their metastases revealed similar 2-DE profiles. In contrast, variations between different lesions with comparable histological characteristics were considerable. Greater differences in polypeptide expression were observed between potentially highly malignant carcinomas compared with comparisons of less malignant lesions. Our results show that malignant human breast carcinomas may be highly heterogeneous in their patterns of gene expression.

Adult↗

PubMed1996

Inhibitory properties coexist with plasticity in the song system of the canary.

The expression of neuronal growth regulating factors (Ni 35/250, GAP-43) were studied immunohistochemically in the plastic telencephalic song system and in the cerebellum of the canary. In the myelinated song nuclei (HVc and RA) growth promoting and inhibiting molecules could be detected, whereas in the white matter tracts of the cerebellum only the neurite growth inhibitor could be detected. The morphodynamic effects of these molecules were assessed by in vitro assays on cryosections. Neurites were formed by 20-30% of the E7 chick cortical neuroblasts cultured in myelinated areas of the vocal control system, neurite outgrowth from the white matter of the cerebellum was completely inhibited.

Animals↗

PubMed1996

Different mechanisms are responsible for c-jun mRNA induction by cisplatin and ultraviolet light.

Ultraviolet light (UV) and different DNA-damaging agents are known to induce AP-l-transcription-factor activity. Whereas UV induction appears to be triggered by events at the cell membrane, the mechanism of AP-l activation by alkylating or platinating agents is not known. We have here examined the effect of cisplatin on AP-l activity in RPMI-8322 melanoma cells. Cisplatin was found to induce binding of nuclear proteins to TRE elements from the c-jun and collagenase-gene promoters, and was also found to induce activation of a c-jun-promoter reporter construct. Compared with stimulation by UV, cisplatin stimulation of c-jun-promoter activity was found to be less sensitive to a dominant negative mutant of Raf-I protein kinase. Furthermore, whereas UV treatment resulted in strong MAP-kinase activation, cisplatin treatment resulted only in a weak and transient increase. These data suggest that the Raf-MAPK pathway is of minor importance for the induction of c-jun-promoter activity by cisplatin. Finally, we report that cisplatin induction of c-jun in RPMI-8322 cells was blocked by herbimycin A, an inhibitor of Src-family tyrosine kinases. In contrast, UV induction of c-jun was not blocked by herbimycin A. In conclusion, our data strongly suggest that UV and cisplatin induction of c-jun mRNA in RPMI-8322 melanoma cells occur by distinct mechanisms.

Benzoquinones↗

PubMed1996

H-89 inhibits collagenase induction by phorbol ester through a mechanism that does not involve protein kinase A.

Cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) regulates the activity of growth-factor-induced pathways at the level of cytoplasmic kinases and nuclear transcription factors. We observed that H-89, an inhibitor of PKA, induced mitogen-activated protein (MAP) kinase activity in a 12V-ras-transformed fibroblast cell line. In contrast, H-89 inhibited phorbol-ester-mediated induction of MAP kinase, junB messenger ribonucleic acid (mRNA), and collagenase mRNA in these cells. Phorbol-ester stimulation of a collagenase-promoter reporter construct was also inhibited by H-89. However, stimulation of the collagenase promoter was not inhibited by overexpression of the PKA-inhibitory protein PKI. These data suggest that H-89 inhibits the activity of an enzyme required for phorbol-ester induction of collagenase mRNA, but that this inhibition does not occur at the level of PKA.

Animals↗

PubMed1996

The metastatic phenotype--prognostic implications.

Strong efforts are being made in order to better understand the molecular mechanisms underlying cancer dissemination. We have attempted to summarise some of the findings in this area. A large number of differences in gene expression have been described in metastatic and non-metastatic cells. In the mouse B16 melanoma system, more than 50 different markers have been described. It is likely that many of these differences reflect the same genetic alteration (i.e. a mutation in a regulatory gene alters the expression of a set of co-regulated target genes). One could argue that it is more effective to study mutations in regulatory as opposed to expression of down-stream target genes. However, we feel that proto-oncogenes are less suitable as markers compared to target genes, since it is difficult to screen for mutations at multiple levels in regulatory pathways. In contrast, measuring the expression of a small number of target genes (i.e. one of the targets in Fig. 1), the expression of which are stimulated by upstream regulators, is accomplished more easily. It is anticipated that the future of optimised panels of independent markers will sharpen cancer diagnosis and lead to individualised therapy.

Animals↗

PubMed1996

Expression of tropomyosin isoforms in benign and malignant human breast lesions.

High molecular weight tropomyosins (tms) are commonly down-regulated in fibroblasts transformed by oncogenes. Previous studies have also demonstrated that specific tm isoforms are down-regulated in human breast carcinoma cell lines. We examined tropomyosin isoforms in cells prepared from non-cancerous breast lesions and primary human breast carcinomas. The average level of expression of all three high molecular weight tm isoforms (tm 1-3) in carcinomas was generally found to be less than 25% of that observed in non-cancerous breast lesions. Interestingly, the expression of tm 1 was found to be 1.7-fold higher in primary tumours with metastatic spread to axillary lymph nodes compared with primary tumours with no evidence of metastasis (p<0.05). Similarly, tm 1 expression was higher in two 12V-H-ras transformed fibroblast cell lines capable of experimental metastasis compared with three weakly metastatic cell lines. We conclude from these studies that expression of high molecular weight tm isoforms is low in primary breast carcinomas, and that metastatic tumours express relatively high levels of tm 1.

Breast↗

PubMed1996

Analysis of polypeptide expression in benign and malignant human breast lesions: down-regulation of cytokeratins.

Malignant progression of tumour cells is caused by the accumulation of genetic defects, which when combined will generate a large phenotypic diversity. Simultaneous quantitation of a large number of gene products in tumour cells is desirable, but difficult to achieve. We have here quantitated the levels of a number of abundant polypeptides in human breast carcinoma cells using two-dimensional gel electrophoresis (2-DE; PDQUEST). For this purpose, tumour cells were prepared from the tissue of 17 breast carcinomas. Fibroadenoma tissue was used as reference for benign cells. An increase of the spot density of the PCNA polypeptide was observed in rapidly proliferating tumour cells, confirming the validity of the procedures used. In the set of 24 polypeptide spots with known identity, decreases in cytokeratin and tropomyosin levels were observed. The levels of all cytokeratin forms resolved (CK7, CK8, CK15 and CK18) were significantly lower in carcinomas than in fibroadenomas. The levels of tropomyosin 2 and 3 were lower in carcinomas than in fibroadenomas. In contrast, the levels of some members of the stress protein family (pHSP60, HSP90 and calreticulin) were higher in carcinomas. Furthermore, changes in the expression of lactate dehydrogenase and GT-pi, but not in nm23, were observed. We conclude that simultaneous analysis of multiple polypeptides in human carcinomas can be achieved by 2-DE and may be useful in prognostic studies, and that malignant progression of breast carcinomas results in the decreased expression of cytokeratin polypeptides. This phenomenon must be considered in studies where cytokeratins are used as markers to identify the epithelial cell compartment in breast carcinomas.

Breast Neoplasms↗