Expression and differential regulation of connective tissue growth factor in pancreatic cancer cells.

CTGF is an immediate early growth responsive gene that has been shown to be a downstream mediator of TGFbeta actions in fibroblasts and vascular endothelial cells. In the present study hCTGF was isolated as immediate early target gene of EGF/TGFalpha in human pancreatic cancer cells by suppression hybridization. CTGF transcripts were found in 13/15 pancreatic cancer cell lines incubated with 10% serum. In 3/7 pancreatic cancer cell lines EGF/TGFalpha induced a significant rise of CTGF transcript levels peaking 1-2 h after the start of treatment. TGFbeta increased CTGF transcript levels in 2/7 pancreatic cancer cell lines after 4 h of treatment and this elevation was sustained after 24 h. Only treatment with TGFbeta was accompanied by a parallel induction of collagen type I transcription. 15/19 human pancreatic cancer tissues were shown to overexpress high levels of CTGF transcripts. CTGF transcript levels in pancreatic cancer tissues and nude mouse xenograft tumors showed a good correlation to the degree of fibrosis. In situ hybridization and the nude mouse experiments revealed that in pancreatic cancer tissues, fibroblasts are the predominant site of CTGF transcription, whereas the tumor cells appear to contribute to a lesser extent. We conclude that CTGF may be of paramount importance for the development of the characteristic desmoplastic reaction in pancreatic cancer tissues.

Cloning of a gene highly overexpressed in cancer coding for a novel KH-domain containing protein.

In a previous large scale screen for differentially expressed genes in pancreatic cancer, we identified a gene highly overexpressed in cancer encoding a novel protein with four K-homologous (KH) domains. KH-domains are found in a subset of RNA-binding proteins, including pre-mRNA-binding (hnRNP) K protein and the fragile X mental retardation gene product (FMR1). By fluorescence in situ hybridization (FISH) the identified gene named koc (KH domain containing protein overexpressed in cancer) was assigned to chromosome 7p11.5. Two pseudogenes were localised on chromosome 6 and 11. The cloned koc cDNA has a 250 bp 5'-UTR, a 1740 bp ORF and a 2168 bp 3'-UTR. The AU-rich 3'-untranslated region of koc contains eight AUUUA and four AUUUUUA reiterated motifs. The deduced koc protein with 580 amino-acids has a relative molecular mass (Mr) of approximately 65,000 (65 K). The koc transcript is highly overexpressed in pancreatic cancer cell lines and in pancreatic cancer tissue as compared to both, normal pancreas and chronic pancreatitis tissue. High levels of expression were as well found in tissue samples of other human tumours. As the KH domain has been shown to be involved in the regulation of RNA synthesis and metabolism, we speculate that koc may assume a role in the regulation of tumour cell proliferation by interfering with transcriptional and or posttranscriptional processes. However, the precise role of koc in human tumour cells is unknown and remains to be elucidated.

Expression of the highly conserved RNA binding protein KOC in embryogenesis.

The human KOC gene which is highly expressed in cancer shows typical structural features of an RNA binding protein. We analyzed the temporal and spatial expression pattern of KOC in mouse embryos at different gestational ages. The expression of KOC seems to be ubiquitous at early stages. During advanced gestation highest KOC expression occurs in the gut, pancreas, kidney, and in the developing brain. The expression pattern of KOC was compared to its Xenopus homologue Vg1-RBP during frog development. Similar expression was found in these organs suggesting an important functional role of the homologous proteins in embryonic development.

Identification of a new tumour-associated antigen TM4SF5 and its expression in human cancer.

In a previous large-scale screening for differentially expressed genes in pancreatic cancer, a gene was identified that was highly overexpressed in pancreatic cancer encoding a novel putative tetraspan transmembrane protein highly homologous to the tumour-associated antigen L6. Using a radiation hybrid panel the identified human gene named TM4SF5 (transmembrane 4 superfamily member 5) was localized to chromosome 17 in the region 17p13.3. The cloned TM4SF5 cDNA has a 32bp 5'-untranslated region (UTR), a 591bp open reading frame (ORF) and a 85bp 3'UTR. The predicted TM4SF5 protein with 197 amino acids contains three NH2-terminal hydrophobic transmembrane regions, followed by an extracellular hydrophilic domain containing two potential N-linked glycosylation sites and a COOH-terminal hydrophobic transmembrane region. These structural features are shared by the L6 antigen and a number of related cell surface proteins associated with cell growth. TM4SF5 was overexpressed in pancreatic cancer tissues as compared to both normal pancreas and chronic pancreatitis tissues, and was detected at high levels in other tumour tissues. Although the precise function of TM4SF5 remains to be elucidated it may be useful in a clinical setting for tumour diagnosis and/or therapy. This hypothesis is supported by the strong homology to the L6 antigen, which has proved promising in immunological, therapeutic and diagnostic approaches.

Depression and reduced natural killer cytotoxicity: a longitudinal study of depressed patients and control subjects.

Cross-sectional studies have demonstrated that natural killer (NK) cell activity is reduced in depression. To extend these observations and examine further the association between severity of depressive symptoms and values of NK activity, this study used a longitudinal case-control design and assessed NK cytotoxicity at intake and at follow-up 6 months after discharge from the hospital in depressed patients and control subjects. From acute hospitalization to follow-up, depression scores significantly (P < 0.01) decreased following treatment in the depressed patients but did not change in the control subjects. NK activity significantly (P < 0.05) increased from intake to follow-up in the depressives while lytic activity did not change in the controls. At intake NK activity was significantly (P < 0.01) reduced in the depressed patients as compared to values in the controls, while at follow-up cytotoxicity was similar between the two groups. These longitudinal data suggest that a reduction of NK cytotoxicity is temporally associated with the state of acute depression.

[Diagnosis of a "hereditary pancreatitis" by the detection of a mutation in the cationic trypsinogen gene]. / Diagnose einer "hereditären Pankreatitis" durch Nachweis der Mutation im kationischen Trypsinogen-Gen.

HISTORY AND CLINICAL FINDINGS: A 71-year-old woman was admitted with the suspected diagnosis of pancreatic carcinoma. As a child she had had repeated attacks of abdominal pain of undetermined cause. When aged 48 years she had developed diabetes mellitus. Her now 42-year-old daughter had from the age of 9 years suffered from repeated attacks of acute pancreatitis that had finally led to chronic pancreatitis. The patient's 15-year-old grandchild was having recurrent bouts of abdominal pain. INVESTIGATIONS: Imaging procedures revealed calcifications in the pancreas and an infiltrating space-occupying lesion, about 3 cm in diameter, in the head of the pancreas with lymph node and liver metastases. Cytological analysis of material aspirated from the space-occupying mass showed typical findings of ductal pancreatic carcinoma. FURTHER TESTS, TREATMENT AND COURSE: At first the patient's course was not typical for a genetically-determined disease, but the family history raised the suspicion of hereditary pancreatitis. A genetic test (Afl-III-RFLP test) demonstrated the mutation Arg 117 His in the cationic trypsinogen gene in all diseased or symptomatic family members. The patient died of the complications of the pancreatic cancer. CONCLUSION: Genetic tests are valuable in the diagnosis of hereditary pancreatitis, because the increased cancer risk can be met by frequent examinations in affected family members.

[Update on the latex allergy topic]. / Aktuelles zum Thema Latex-Allergie.

Type I allergies to latex have become an increasing problem in occupational dermatology during the past few years, especially since at least 10% of health care workers are affected. In the Department of Dermatology, University Erlangen-Nuremberg, a 12-fold increase in latex-allergic patients has been documented between 1989 and 1995 with a clear trend to more severe systemic manifestations (from 10.7% in 1989/ 1990 to 44% in 1994/1995). Among the water soluble proteins (molecular weights 2 to 200 kD) which may induce latex allergy, at least 5 are considered as main proteins with known primary structure. In addition some "marker' proteins seem to induce specific IgE antibodies in special risk groups (e.g. 46 kD-protein in medical professions, 14.6 kD- and 27 kD-proteins in children with spina bifida). Cross reactions between latex and several fruits (especially avocado, kiwi, banana and chestnut) in 60 to 70% of latex-allergic patients have to be taken into account when evaluating and counselling affected patients. Most important in prophylaxis is the complete change to powder-free latex gloves in medical institutions, since these gloves usually have a low protein content. Our listing of surgical and examination gloves according to their protein content (as measured by the modified Lowry- and High Pressure Liquid Chromatography method) should be a useful guideline for the choice of suitable gloves.

KOC is a novel molecular indicator of malignancy.

The detection of malignant cells in fine-needle aspirates (FNA's) using marker genes is hampered by the fact that these markers are only expressed by certain malignancies or lack sensitivity and/or specificity. Here we report the results of a prospective pilot study examining the expression of KOC (KH-domain containing protein over expressed in cancer), a novel onco-foetal gene, in 76 patients who underwent fine-needle aspiration for further diagnosis of abdominal lesions, aszites, cysts or cerebrospinal fluid. Aspirates were examined by cytology and by a KOC RT-PCR assay. KOC expression was a highly sensitive and specific indicator of malignancy. The KOC assay could be useful to facilitate screening for malignant disease and to improve the diagnostic accuracy of FNAs.

Identification of genes with specific expression in pancreatic cancer by cDNA representational difference analysis.

cDNA representational difference analysis (cDNA-RDA) is a polymerase-chain-reaction-coupled subtractive and kinetic enrichment procedure for the isolation of differentially expressed genes. In this study, the technique was used to isolate novel genes specifically expressed in pancreatic cancer. cDNA-RDA was done on cDNA reverse transcribed from a poly(A)+ mRNA pool made from 10 cancer tissues (tester) by using as a driver a cDNA from a poly(A)+ mRNA pool made from a combination of 10 tissues of chronic pancreatitis and 10 healthy pancreatic tissues. The use of chronic pancreatitis in addition to healthy pancreas mRNA in the driver preparation eliminated the influence of stromal tissue components present as contamination in the cancer-specific preparations. Such cDNA-RDA led to the isolation of 16 distinct, cancer-specific gene fragments. These were confirmed to be overexpressed in pancreatic cancer tissues by Northern blot analysis. Sequence analysis revealed homologies to five genes previously implicated in the carcinogenesis of the pancreas or other tissues. Eleven fragments had no significant homology to any known gene and thus represent novel candidate disease genes. The experiments demonstrate that cDNA-RDA is a reproducible and highly efficient method for the identification of novel genes with cancer-specific expression.

Cloning of novel transcripts of the human guanine-nucleotide-exchange factor Mss4: in situ chromosomal mapping and expression in pancreatic cancer.

In a previous large-scale analysis of gene expression in pancreatic cancer using gridded arrays of cDNA libraries and differential hybridizations, a gene that was a homolog to human mss4 was identified. Mss4 is a guanine-nucleotide-exchange factor for the Sec4/ Ypt1/Rab family of small GTP-binding proteins involved in the regulation of intracellular vesicular transport. By fluorescence in situ hybridization the human mss4 gene was assigned to chromosome 1q32-q41. Northern blot analysis revealed that three mss4 mRNA species are transcribed in human tissues of 780, 1200, and 2800 bp in length, respectively. Cloning and sequencing of the human mss4 transcripts from a pancreatic cancer cDNA library revealed that these mRNA species differ in the length of the 3-untranslated region and are probably due to the alternate use of polyadenylation sites. All mRNA species were detected at moderate to high levels in pancreatic cancer cell lines and were overexpressed in pancreatic cancer tissue compared to both normal pancreas and chronic pancreatitis tissue. However, the 1200-bp transcript was the Mss4 mRNA species with the highest level of expression in more than 50% of tumor cells and tissues. High levels of expression were found as well in other human tumor tissues. Mss4 as guanine-exchange factor required in the regulation of intracellular transport may be of importance for the function and growth of human tumor cells. However, the precise role of mss4 in human tumor cells is unknown and remains to be elucidated.

Role of MT-MMPs and MMP-2 in pancreatic cancer progression.

Activation of matrix metalloproteinase-2 (MMP-2) by the membrane-type matrix metalloproteinases (MT-MMPs) has been associated with tumor progression. In the present study, we examined the role of MMP-2 and its activators MT1-MMP, MT2-MMP and MT3-MMP in pancreatic tumor cell invasion and the development of the desmoplastic reaction characteristic of pancreatic cancer tissues. Northern blot analyses revealed that transcript levels of MT1-MMP and MT2-MMP, but not MT3-MMP, were enhanced in pancreatic cancer tissues (n = 18) compared with both chronic pancreatitis (n = 9) and healthy pancreas (n = 9). A good correlation was found between MT1-MMP and both MMP-2 expression (p < 0.01) and activity in pancreatic cancer tissues. In addition, expression and activation of MMP-2 were strongly associated with the extent of the desmoplastic reaction in pancreatic cancer tissues. Invasion assays showed a good correlation between MMP-2 expression and activity and the invasive potential of pancreatic cancer cell lines. In cell lines with high levels of MMP-2 expression and activity, the MMP inhibitor Batimastat led to significant reduction of the number of invading cells. Our results suggest that MT1-MMP is involved in the progression of pancreatic cancer via activation of MMP-2. MMP-2 itself plays an important role in tumor cell invasion and appears to be associated with the development of the characteristic desmoplastic reaction in pancreatic cancer.