Proangiogenic potency of MUC18 in arteriosclerotic disease.

AIM: Stenotic peripheral and dilatative arteriosclerotic diseases have different pathomechanism although associations between both diseases are well known. The adhesion molecule MUC18 is a cell membrane glycoprotein also known as the melanoma cell adhesion molecule. As MUC18 has proangiogenic potency in melanoma and prostate cancer this study investigated the role of MUC18 in patients with stenotic or dilatative arteriosclerotic disease as a putative biochemical marker. METHODS: Using qRT-PCR, Western Blot and immunohistochemistry techniques, the expression of MUC18 in arteriosclerotic arteries from major lower limb amputates (AP, N.=15) as well as specimen from femoral endarterectomies (TEA, N.=20) and in dilatative aortic diseases using abdominal aortic aneurysms (AAA, N.=13) was evaluated. Human visceral arteries without macroscopic arteriosclerosis from liver transplants served as controls (AN, N.=19). RESULTS: MUC18 mRNA and protein expression could be found in AN, AP, TEA and AAA tissues. Immunohistochemical analysis showed that a complete and intact intima was the predominant location of MUC18 expression. Although in stenotic arteriosclerotic disease (AP and TEA) the intima was widely calcified, qRT-PCR analysis showed overexpression compared to normal tissue. Interestingly, MUC18 expression was significantly down-regulated in dilatative compared to stenotic arteriosclerotic disease and normal arteries. CONCLUSION: In peripheral stenotic arteriosclerotic disease the proangiogenic potency of MUC18 may play a role in angiogenesis of collaterals, whereas in dilatative aortic diseases the induction of collaterals is typically not evident. The results support the hypothesis of a role in angiogenesis of MUC18 in stenotic arteriosclerotic disease.

CCR6/CCL20 chemokine expression profile in distinct colorectal malignancies.

Originally, chemokines and their G-protein-coupled receptors were described to regulate multiple physiological functions, particularly tissue architecture and compartment-specific migration of white blood cells. Now, it is established that the chemokine/chemokine receptor system is also used by cancer cells for migration and metastatic spread. Here, we examined the relative levels of CC-chemokine CCL20 and its corresponding receptor CCR6 in resection specimens from patients with different malignant and non-malignant colorectal diseases as well as in colorectal liver metastases (CRLM). CCL20/CCR6 mRNA and protein expression profiles were assessed by quantitative real-time PCR (qRT-PCR), enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry (IHC) in resection specimens from patients with ulcerative colitis (UC, n = 15), colorectal adenoma (CRA, n = 15), colorectal adenocarcinoma (CRC, n = 61) and colorectal liver metastases (CRLM, n = 16). Corresponding non-diseased tissues served as control. In contrast to UC tissues, the CCL20/CCR6 system showed a distinct upregulation in CRA, CRC and CRLM related to corresponding non-affected tissues (P < 0.05, respectively). Furthermore, CRA, CRC and CRLM tissue samples displayed significantly higher protein amounts of CCL20 in comparison with UC specimens (P < 0.05, respectively). Our results strongly suggest an association between CCL20/CCR6 expression and the induction of CRA, CRC and the development of CRLM. Therefore, CCL20 and CCR6 may provide potential targets for novel treatment strategies of CRC.

Differential CXC receptor expression in colorectal carcinomas.

In this study, we aimed to assess the expression profile of chemokine receptors CXCR1-4 in inflammatory and malignant colorectal diseases and corresponding hepatic metastases of synchronous and metachronous origin to elucidate their role in colorectal cancer (CRC) progression and metastasis. Chemokine receptor expression was assessed by quantitative real-time PCR, immunohistochemistry (IHC) and Western blot analysis in resection specimens from patients with ulcerative colitis (UC, n = 25), colorectal adenomas (CRA, n = 8), different stages of CRC (n = 48) as well as colorectal liver metastases (CRLM) along with their corresponding primary colorectal tumours (n = 16). While none of the chemokine receptors were significantly upregulated or downregulated in UC or CRA tissues, CXC receptors 1, 2 and 4 demonstrated a significant increase in expression in all tumour stages of CRC specimens with CXCR4 correlating with tumour grading (P < 0.05). On the other hand, CXCR3 showed no significant upregulation in either tumour stage, but significant overexpression in CRLM. While CXCR4 demonstrated significant upregulation in both tumour entities, IHC analysis revealed that the predominate cell type expressing CXCR4 in CRC is represented by tumour cells, whereas in CRLM the majority of positive CXCR4 signals is due to hepatocytes along the tumour invasion front. In conclusion, our findings show a very differential expression pattern of the four receptors in colorectal carcinomas and their corresponding liver metastases with prominent expression profiles that indicate a potential role in the pathogenesis of CRC.

Enhanced expression and clinical significance of CC-chemokine MIP-3 alpha in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most frequent visceral neoplasms worldwide. Using RT-PCR, ELISA, microdissection and immunohistochemistry, we investigated the expression profiles of CCL19, CCL20, CCL21 and CXCL12 and their receptors in tumourous and tumour neighbouring tissues from patients with HCC and in nonmalignant liver lesions, respectively. All chemokines were found to be expressed in normal liver and HCC tissues, yet CCL20 was the only chemokine showing significant upregulation in HCC tissues. Clinicopathological analysis revealed a distinct increase in CCL20 expression rates in HCC tissues of grade III tumours in comparison to HCC tissues from grade II tumours. On mRNA level, only chemokine receptor CCR6 revealed significant upregulation in HCC tissues. However, immunohistochemical studies indicated a marked CCR6 expression accumulated in a streak of normal cells along the tumour invasion front in all our HCC specimens which could provide a stimulative signal for the tumour to further expand. The present findings show significant overexpression of CCL20 in the tumour tissues and marked overexpression of the corresponding receptor CCR6 in the tumour invasion front of HCC patients in comparison to normal liver. Moreover, CCL20 expression was found to correlate with tumour grade and therefore, we suggest that the CCL20/CCR6 system may be involved in hepatocarcinogenesis.

Mutation analysis of the zinc transporter gene SLC30A4 reveals no association with periodic catatonia on chromosome 15q15.

The zinc transporter gene SLC30A4, located on chromosome 15q15-q21, has previously been reported to show altered expression patterns in post mortem analysis of the brains of schizophrenic patients. As a positional candidate we investigated SLC30A4 in the chromosome 15q15-linked schizophrenic phenotype periodic catatonia (MIM 605419), by means of a systematic mutation screening in affected individuals from exceptionally large pedigrees with perfect co-segregation of a chromosomal segment between marker D15S1042 and D15S659 in all affected individuals. The mutation scan revealed no genetic variants within the coding and the putative promoter region of SLC30A4 and, thus, excludes a genetic association of SLC30A4 with catatonic schizophrenia.

Reduced amplification efficiency of KIAA0027/MLC1 alleles: implications for the molecular diagnosis of megalencephalic leukoencephalopathy with subcortical cysts.

Autosomal recessive megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a rare childhood-onset spongiform leukodystrophy with macrocephaly and slowly progressive deterioration of motor functions. Mutations in KIAA0027/MLC1 have recently been found associated with MLC, and a high degree of allelic heterogeneity has been observed. In addition, initial reports suggested that a rare variant in exon 11 (L309M) is involved in the etiology of schizophrenia, but recent studies have brought forward compelling arguments that genetic variants of MLC1 are not associated with schizophrenia. Using DHPLC-analysis, reproduction of previous findings on L309M revealed homoduplex resolution patterns among individuals, who had been described to be heterozygous for the variant, which was further confirmed by sequencing the respective PCR products. Cumulative effects of high GC content, secondary folding structures due to incomplete intronic tandem-repeats, and a complicated insertion polymorphism at the 3-end of exon 11 may be the cause of preferential amplification of specific alleles of exon 11. Consistent amplification was obtained only when we employed exonic primers directly adjacent to the L309M variant. For mutational screening, we propose a two-step test: (1) testing for the 33 bp insertion polymorphism of exon 11, and (2) amplification of the exon using different primer sets depending on the presence or absence of the insertion.

The human dopamine transporter gene: the 5'-flanking region reveals five diallelic polymorphic sites in a Caucasian population sample.

The 5'-flanking region of the human dopamine transporter (hDAT) was systematically screened for variants by single strand conformation analysis (SSCA) between -1586 and +97 basepair (bp) relative to the transcription start site. Five diallelic polymorphisms were found, which were shown to be due to single base substitutions: T-67A, G-660C, C-839T, C-1169G, T-1476G. In a population sample of 119 unrelated Caucasians, allele frequencies of the rarer allele were 47% for -67T, 3% for -660C, 45% for -839T, 50% for -1169G, and 8% for -1476G, respectively. Among 15 observed haplotypes, seven haplotypes collected a frequency of about 96% in our sample. T-67A, C-839T, C-1169G, T-1476G were related to potential transcriptional recognition sites. These findings and the occurrence of distinct haplotypes at the hDAT promoter locus in a Caucasian population sample make this region a promising target in the context of linkage and association studies in certain diseases.

Autosomal recessive long-QT syndrome (Jervell Lange-Nielsen syndrome) is genetically heterogeneous.

Jervell Lange-Nielsen syndrome (JLNS) is a recessive disorder with congenital deafness and long-QT syndrome (LQTS 1). Mutations in the potassium-channel gene KVLQT1 (LQTS 1) have been identified in JLNS and in autosomal-dominant LQTS as well. We performed haplotype analysis with microsatellite markers in a Lebanese family with JLNS, but failed to detect linkage at LQTS 1. Moreover, using this approach, we excluded two other ion-channel genes involved in autosomal-dominant LQTS, HERG (LQTS 2) and SCN5A (LQTS 3). Our findings indicate that JLNS is genetically heterogeneous and that, in this family, an unknown LQTS gene causes the disease.

Juvenile myelomonocytic leukemia: analyses of treatment results in the EORTC Children's Leukemia Cooperative Group (CLCG).

Forty-six children with juvenile myelomonocytic leukemia (JMML) diagnosed between 1978 and 1993 in 12 centers were retrospectively studied. There is no evidence that any conventional treatment influences the long-term evolution of JMML. Among 28 patients treated without bone marrow transplantation (BMT), 26 died (median survival: 17 months), two are alive, one in complete remission (CR) after intensive chemotherapy. Allogenic BMT is the best treatment: 18 patients underwent BMT, 11 are in CR (at 9, 15, 22, 25, 41, 45, 49, 53, 66, 90 and 108 months). Conditioning regimens using chemotherapy alone may cure some patients (3/6) occasionally despite autologous reconstitution (1/3); if relapse occurs, a second BMT may be curative (2/3). Among the 12 patients conditioned immediately with TBI, six are in CR, one is in relapse, five died (one of them in durable autologus CR from Schwannoma). It is our opinion that splenectomy is of therapeutic value and seems not to have influenced the incidence of infections complications. We found no argument in favor of intensive chemotherapy before conditioning. Results with HLA-matched unrelated donors are satisfactory. One patient relapsed at 4 months after an unrelated BMT and entered a new CR after discontinuation of cyclosporine.