Martina Zeljko

Brain Metastases from Lung Cancer Show Increased Expression of DVL1, DVL3 and Beta-Catenin and Down-Regulation of E-Cadherin

The susceptibility of brain to secondary formation from lung cancer primaries is a well-known phenomenon. In contrast, the molecular basis for invasion and metastasis to the brain is largely unknown. In the present study, 31 brain metastases that originated from primary lung carcinomas were analyzed regarding over expression of Dishevelled-1 (DVL1), Dishevelled-3 (DVL3), E-cadherin (CDH1) and beta-catenin (CTNNB1). Protein expressions and localizations were analyzed by immunohistochemistry. Genetic alterations of E-cadherin were tested by polymerase chain reaction (PCR)/loss of heterozygosity (LOH). Heteroduplex was used to investigate mutations in beta-catenin. DVL1 and DVL3 showed over expression in brain metastasis in 87.1% and 90.3% of samples respectively. Nuclear staining was observed in 54.8% of cases for DVL1 and 53.3% for DVL3. The main effector of the Wnt signaling, beta-catenin, was up-regulated in 56%, and transferred to the nucleus in 36% of metastases. When DVL1 and DVL3 were up-regulated the number of cases with nuclear beta-catenin significantly increased (p = 0.0001). Down-regulation of E-cadherin was observed in 80% of samples. Genetic analysis showed 36% of samples with LOH of the CDH1. In comparison to other lung cancer pathologies, the diagnoses adenocarcinoma and small cell lung cancer (SCLC) were significantly associated to CDH1 LOH (p = 0.001). Microsatellite instability was detected in one metastasis from adenocarcinoma. Exon 3 of beta-catenin was not targeted. Altered expression of Dishevelled-1, Dishevelled-3, E-cadherin and beta-catenin were present in brain metastases which indicates that Wnt signaling is important and may contribute to better understanding of genetic profile conditioning lung cancer metastasis to the brain.

Changes of AXIN-1 and Beta-Catenin in Neuroepithelial Brain Tumors

In the present study changes of components of Wnt signaling pathway—axin (AXIN1) and beta-catenin (CTNNB1) in a sample of 72 neuroepithelial brain tumors were investigated. AXIN-1 gene was tested by PCR/loss of heterozygosity (LOH). Immunostaining and image analysis revealed the quantity and localization of relevant proteins. Polymorphic marker for AXIN-1, showed LOH in 11.1% of tumors. LOH was distributed to 6.3% of glioblastomas, one was found in neuroepithelial dysembrioplastic tumor and one in medulloblastoma. Down regulation of axin expression and up regulation of beta-catenin were detected in the analyzed tumors. Axin was observed in the cytoplasm in 68.8% of samples, in 28.1% in both the cytoplasm and nucleus and 3.1% had no expression. Beta-catenin was observed mainly in the nucleus and cytoplasm (59.4%). Expression in 34.4% of samples was in the cytoplasm and 6.3% showed no expression. Comparison of mean values of relative increase of axin and beta-catenin showed that they are significantly reversely proportional (P = 0.014). Relative quantity of beta-catenin in patients with gross deletion of AXIN1 was significantly higher in comparison to patients without LOH (P = 0.040). Our results demonstrate that changes of key components of the Wnt signaling play a role in neuroepithelial brain tumors.

Report on mutation in exon 15 of the APC gene in a case of brain metastasis

The study analyzes exon 15 of the adenomatous polyposis coli gene (APC) in a 49-year-old male patient with brain metastasis. The primary site was lung carcinoma. PCR method and direct DNA sequencing of the metastasis and autologous lymphocyte samples identified the presence of a somatic mutation. The substitution was at position 5883 G–A in the metastasis tissue. The mutation was confirmed by RFLP analysis using Msp I endonuclease, since the mutation strikes the Msp I restriction site. Immunohistochemical analysis revealed the lack of protein expression of this tumor suppressor gene. The main molecular activator of the wnt pathway, beta-catenin, was expressed, and located in the nucleus. The mutation is a silent mutation that might have consequences in the creation of a new splice site. Different single-base substitutions in APC exons need not only be evaluated by the predicted change in amino acid sequence, but rather at the nucleotide level itself. In our opinion, such silent mutations should also be incorporated in mutation detection rate and validation.

Expression of Secreted Frizzled-Related Protein 1 and 3, T-cell Factor 1 and Lymphoid Enhancer Factor 1 in Clear Cell Renal Cell Carcinoma

Frequency and mortality of renal cell carcinoma (RCC) are increasing for decades. However, the molecular background of RCC tumorigenesis is still poorly understood. In current study we investigated the expression of TCF/LEF and SFRP family members (SFRP1 and SFRP3) to gain a better understanding of biological signaling pathways responsible for epidemiology and clinical parameters of clear cell RCC (cRCC). Thirty-six pairs of paraffin-embedded clear cRCC and adjacent nontumoral tissues samples using immunohistochemistry (IHC) were analyzed and compared with corresponding clinicopathological parameters. Immunohistochemistry indicated statistically significant decreased SFRP3 expression in tumor tissues but no consistency in SFRP1 expression in analyzed normal and tumor tissue. The TCF1 expression level was significantly weaker in normal tissue compared to tumor samples while LEF1 protein levels were significantly weaker in tumor tissue. To our knowledge, this is the first report on analysis of the expression of transcription factors TCF1 and LEF1 in clear cell renal cell carcinoma and their comparison with Wnt signal pathway antagonists belonging to SFRP family.

Brain metastases exhibit gross deletions of the APC gene

Candidate genes involved in metastasis to the brain require investigation. In the present study, the adenomatous polyposis coli (APC) gene was analyzed in a set of human brain metastases. Gross deletions of the APC gene were tested by polymerase chain reaction/loss of heterozygosity (LOH) using the restriction fragment length polymorphism method performed by the use of MspI and RsaI genetic markers inside exon 15 and exon 11. Among 21 brain metastases analyzed, 58.8% of samples showed LOH of the APC gene. When assigning the genetic changes to a specific primary tumor type, 6 LOHs were found in metastases originated from lung and 4 LOHs in metastases from colon. The main effector of the wnt signaling, beta-catenin, was upregulated in 42.9% of cases and transferred to the nucleus in 28.6% of metastasis cases. Our findings suggest that genetic changes of the tumor suppressor gene APC, a component of the wnt pathway, represent a part of the brain metastasis genetic profile.

Frequency of loss of heterozygosity of the NF2 gene in schwannomas from Croatian patients

Aim To identify gross deletions in the NF2 gene in a panel of schwannomas from Croatian patients in order to establish their frequencies in Croatian population. Methods Changes of the NF2 gene were tested by polymerase chain reaction/loss of heterozygosity (LOH) using two microsatellite markers, D22S444 and D22S929. Results The analysis with both markers demonstrated that 43.75% of schwannomas exhibited LOH of the NF2 gene. The D22S444 region exhibited 45.5% of LOHs and the D22S929 region exhibited 14.3% of LOHs. Four LOHs were found in Antoni B, 2 in Antoni A, and 1 in Antoni A and B type tumors. Conclusion The frequency of changes observed in Croatian patients is broadly similar to that reported in other populations and thus confirms the existing hypothesis regarding the tumorigenesis of schwannomas and contributes to schwannoma genetic profile helping us to better understand its etiology and treatment.

Utjecaj modifikacije višestjenih ugljikovih nanocjevčica na svojstva poliuretana: II. Mehanička svojstva, električna provodnost i toplinska postojanost

Za razliku od jednostjenih ugljikovih nanocjevčica (SWCNT) koje se sastoje od jednog grafenskog sloja smotanog u cilindar, višestjene ugljikove nanocjevčice (MWCNT) izgrađene su od više smotanih grafenskih slojeva. Ugljikove nanocjevčice, zbog iznimnih mehaničkih, toplinskih i električnih svojstava, predstavljaju skupinu nanopunila koja mogu znatno poboljšati svojstva polimera.2,3 Međutim zbog nanodimenzija i van der Waalsovih međudjelovanja između pojedinačnih ugljikovih nanocjevčica izrazito su sklone agregiranju,2 što predstavlja problem u postizanju jednolike raspodjele u polimernoj matrici. Modifikacija površine ugljikovih nanocjevčica predstavlja značajan smjer u savladavanju tog problema.4

Utjecaj modifikacije višestjenih ugljikovih nanocjevčica na svojstva poliuretana: I. Morfologija i toplinska svojstva

Poliuretanski elastomeri su blok-kopolimeri, koji se sastoje od krutog i fleksibilnog segmenta. Zbog razlike u polarnosti odnosno termodinamičke nekompatibilnosti, ti segmenti se razdvajaju u meku i tvrdu fazu. Zahvaljujući strukturi, poliuretanski elastomeri imaju superiorna svojstva povezana s visokom tvrdoćom s obzirom na modul, visokom otpornošću na abraziju, izvrsnim mehaničkim svojstvima te biokompatibilnosti i stoga imaju iznimno široku primjenu u području adheziva, optoelektronike, biotehnologije i brojnim drugim.1,2