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Biomedical subjects

J Pavelić

Publications and source records attributed to J Pavelić.

At least 19 recordsLinked to original sources
PubMed2002

Novel alleles of the D16S752 polymorphic genetic marker linked to E-cadherin gene--a potential population marker.

Seven DNA variants that polymorphic genetic marker D16S752 reveals in Croatian population are reported in this paper. The marker is a GATA tetranucleotide repeat linked to human E-cadherin gene (CDH1). Prior studies involving this marker revealed only four DNA allele variants. The reported DNA variants contribute to the collection of hypervariable DNA polymorphisms data useful in the field of anthropological and population genetic and forensic medicine.

Cadherins↗
PubMed2001

The efficacy of retroviral herpes simplex virus thymidine kinase gene transfer and ganciclovir treatment on the inhibition of melanoma growth in vitro and in vivo.

One approach to gene therapy of cancer is based on the insertion of a suicide gene into tumor cells and subsequent activation of the suicide mechanism. We used the herpes simplex virus thymidine kinase (HSVtk) gene followed by ganciclovir (GCV) treatment. The goal of our experiments was to determine the effectiveness of HSVtk gene therapy in malignant melanoma. B16BL6 murine melanoma cells retrovirally transduced with the HSVtk gene became sensitive to low concentrations of GCV. Analysis by RT-PCR showed HSVtk expression in transduced B16BL6tk+ cells. Apoptotic cell death was found in B16BL6tk+ cells treated with GCV (20 microM). The sensitivity of B16BL6tk+ cells to GCV was also examined in vivo. Tumors inoculated subcutaneously into C57BL6 mice regressed rapidly when treated with GCV (50 mg/kg twice a day) and disappeared completely after 14 days treatment. The mice remained in remission for 5 months. A bystander effect through which nontransduced B16BL6 cells were also inhibited by GCV administration when cocultured with B16BL6tk+ cells was expected. However, only slight killing of nontransduced cells was observed in vitro. Analysis of the bystander effect in vivo showed complete regression of tumors inoculated with a mixture of cells mostly consisting of B16BL6tk+ cells. A distant bystander effect was also examined. There was no regression of wild-type tumors raised at a distant site from primary B16BL6tk+ tumors. The failure of a more effective bystander effect indicates the need for further investigation of the possible use of combined gene therapy to treat melanoma.

3T3 Cells↗
PubMed2001

Natural zeolite clinoptilolite: new adjuvant in anticancer therapy.

Natural silicate materials, including zeolite clinoptilolite, have been shown to exhibit diverse biological activities and have been used successfully as a vaccine adjuvant and for the treatment of diarrhea. We report a novel use of finely ground clinoptilolite as a potential adjuvant in anticancer therapy. Clinoptilolite treatment of mice and dogs suffering from a variety of tumor types led to improvement in the overall health status, prolongation of life-span, and decrease in tumors size. Local application of clinoptilolite to skin cancers of dogs effectively reduced tumor formation and growth. In addition, toxicology studies on mice and rats demonstrated that the treatment does not have negative effects. In vitro tissue culture studies showed that finely ground clinoptilolite inhibits protein kinase B (c-Akt), induces expression of p21WAF1/CIP1 and p27KIP1 tumor suppressor proteins, and blocks cell growth in several cancer cell lines. These data indicate that clinoptilolite treatment might affect cancer growth by attenuating survival signals and inducing tumor suppressor genes in treated cells.

Adjuvants, Pharmaceutic↗
PubMed2001

Helicobacter pylori colonization of tongue mucosa--increased incidence in atrophic glossitis and burning mouth syndrome (BMS).

Nested polymerase chain reaction (PCR) was performed to detect the presence of Helicobacter pylori in tongue mucosa in 268 patients divided into four groups according to their diagnosis: 87 with atrophic glossitis, 37 with benign migratory glossitis and 144 with burning mouth syndrome (BMS). The latter group was subdivided according to anatomic site of burning sensation: subgroup A (54 patients) with complaints limited to tongue and subgroup B (90 patients) with burning sensations in other parts of oral mucosa. H. pylori was found in 43 samples (16%). Bacteria were significantly less present in tongue mucosa affected with benign migratory glossitis compared with atrophic glossitis and BMS (P=0.025). This difference was more obvious when compared with atrophic glossitis only (P=0.006). Mucosal changes in these conditions might make the oral environment more acceptable for H. pylori colonization compared with normal mucosa, and this mechanism may play a role in its oro-oral transmission.

Burning Mouth Syndrome↗
PubMed1999

Loss of heterozygosity and protein expression of APC gene in renal cell carcinomas.

This study evaluated the potential contribution of the APC gene to malignant transformation in patients with renal cell carcinoma. We tested 36 human renal cell carcinoma samples and 18 adjacent normal kidney tissues for the expression of APC protein, both wild and truncated types, by western blot using antibodies that recognize either the carboxy or the amino epitope of the APC protein. The same tumor samples together with autologous peripheral blood were also analyzed at the DNA level. Using specific oligonucleotide primers for exons 11 and 15, gene instability was followed by polymerase chain reaction/loss of heterozygosity (LOH) (on the basis of restriction fragment length polymorphism). Molecular data were also compared to pathohistological diagnosis, TNM stage, and patient's age using multivariate statistical methods. All normal renal tissues revealed expression of the wild-type APC protein. Neither wild nor mutant type proteins were found in 36% (13/36) of tumor samples; the rest of tumor tissues expressed the wild-type protein (312 kDa). Mutated APC protein, with a molecular weight of 117 kDa, was found in only one tumor sample. From 36 tumor samples 16 (44.4%) were informative for RsaI exon 11 polymorphic site, while only half of these (8/16) demonstrated LOH. From 13 tumor samples that had no detectable protein product by western blot analysis eight were homozygous for the exon 11 polymorphism and were tested for another polymorphic site, MspI/exon 15. The overall proportion of LOH cases for both polymorphisms tested was 52.9% (9/17). Pathohistological diagnosis and molecular data showed no correlation. However, multivariate analysis determined a stage strong positive correlation of age and TNM with the presence of LOH and the absence of the wild-type APC protein. Out results suggest that the APC tumor suppressor gene plays a role in renal carcinogenesis. Alterations in this gene are responsible for tumor evolution and progression, but cannot be considered as a first event in tumor initiation.

Adenomatous Polyposis Coli Protein↗
PubMed1999

The expression and role of insulin-like growth factor II in malignant hemangiopericytomas.

Hemangiopericytoma is a rare soft tissue tumor originating from contractile pericapillary pericytes. To address the issue of molecular genetic events that participate in genesis and progression of hemangiopericytoma we analyzed insulin-like growth factor (IGF) II and IGF I receptor in 29 tumors collected from a human tumor bank network. Seven of these tumors were associated with severe hypoglycemia; six were retroperitoneal and one was located in the leg. Of 22 tumors tested 12 (54.5%) exhibited IGF II mRNA, while almost 90% (17 of 19) of hemangiopericytomas exhibited IGF I receptor mRNA. Sera from some patients whose tumors expressed IGF II mRNA contained elevated levels of IGF II. Removal of the tumor eliminated most of the IGF II immunoreactivity from the sera. The potential role of IGF II as a growth-promoting factor was examined on three malignant primary hemangiopericytoma cell cultures. Extracellular addition of IGF II significantly enhanced cell proliferation in a dose-dependent manner. Antisense oligodeoxynucleotides that specifically inhibit IGF II mRNA, at a concentration of 40 or 80 micrograms/ml, inhibited the growth of hemangiopericytoma cells significantly, by 40%. Simultaneous administration of antisense deoxyoligonucleotides to both IGF II and IGF I receptor inhibited tumor cell proliferation by even 80%. Our data suggest that tumor cells produce IGF II, and that this in turn stimulates their proliferation by autocrine mechanisms.

Adolescent↗
PubMed1998

Molecular pathology of hemangiopericytomas accompanied by severe hypoglycemia: oncogenes, tumor-suppressor genes and the insulin-like growth factor family.

Relatively little is known about molecular genetic events that participate in the genesis and progression of hemangiopericytoma. In this study, we describe two cases of hemangiopericytoma accompanied by severe hypoglycemia. Tumor cells from patient 1 exhibited insulin-growth factor I (IGF I) and insulin-like growth factor I receptor (IGF IR) mRNA transcripts. Tumor cells from patient 2 exhibited IGF II, IGF IR and IGF binding proteins 1-3 mRNA. Serum from patient 2 contained IGF II, mostly in a large molecular form ("big" IGF II); the IGF II level did not change after the tumor removal. The presence of IGF IR in tumor cells was confirmed by immunoprecipitation with antibodies that recognize human IGF IR subunit (visualized as a 460-kDa band). The hemangiopericytoma cells derived from patient 1 expressed 210000 IGF I receptors/cell. Specific binding of IGF I to the tumor cell membrane fraction was higher in tissue from patient 1, while the tissue of patient 2 showed relatively low IGF I binding. In contrast, IGF II binding was much higher in tissue from patient 2. Both tumor tissues showed positive immunostaining for c-Jun; one tumor showed strong immunostaining for c-Myc, H-Ras and p53, while the other exhibited strong reaction with H-Ras antibodies only. No loss of the heterozygosity at the genes APC, NFI and nm23-H1 loci in tumor tissue obtained from patient 1 was found. In effect, our results suggest multiple molecular genetic changes in hemangiopericytoma -- activation of some oncogenes and the IGF growth factor family. IGF ligands together with IGF IR could be responsible for hypoglycemia and perhaps the transformed phenotype.

Abdominal Neoplasms↗
PubMed1998

Detection of Helicobacter pylori in various oral lesions by nested polymerase chain reaction (PCR).

Nested PCR was used for the detection of Helicobacter pylori DNA in specimens collected from seven different topographic sites in the oral cavity. Out of 161 patients, only 21 (13.04%) were positive. There was no correlation between H. pylori status and patient diagnosis and age. No preferential site for bacterial colonization was found in the oral cavity, nor was an association established between a bacterial presence and ulcerated versus non-ulcerated lesions. The results indicate that the oral mucosa does not appear to represent a preferred site of colonization for H. pylori. Furthermore, the evidence presented in this paper suggests that H. pylori is not pathogenic in the oral cavity, nor is it associated with common oral pathologic processes.

Adolescent↗
PubMed1997

Squamous cell lung carcinomas: the role of nm23-H1 gene.

This analysis of 32 pairs of human squamous cell lung carcinomas and normal matched control DNA demonstrates that loss of heterozygosity (LOH) is infrequent at the nm23-H1 locus, affecting only 2 of the 18 informative cases. Both LOH cases were in the tumor stage IIIA. One tumor was of poor and the other of moderate histological grade. These and an additional 34 tumor samples were also analyzed immunohistochemically for the presence of nm23-H1 protein. Of the 66 cases tested for the presence of nm23-H1 protein 54 were negative. Eight samples exhibited up to 35% positive cells (with weak immunostaining intensity) and four between 35% and 70% (moderate immunostaining intensity); no sample showed more than 70% positive cells. Noncancerous lung parts contained no nm23-H1 protein. nm23-H1 expression was independent of TNM stage, grade, tumor size, and patient's survival. Two samples with LOH were negative for nm23-H1 protein. We therefore conclude that neither loss of heterozygosity of the nm23-H1 gene nor the intensity of specific protein expression are related to squamous cell lung carcinoma development and progression.

Aged↗
PubMed1996

PCR amplification of DNA from archival specimens. A methodological approach.

The experiments were designed to assess whether DNA could be recovered from archival, fixed, paraffin embedded specimens, 1-39 years old, for use in PCR and Southern blot analyses. Specimen fixed in either 10% formalin, Carnoy's or AMeX fixative produced almost equally abundant 318 bp long beta-actin fragment, after 2 x 120 min deparaffinization time, proteinase K DNA extraction and 40, or more, amplification cycles. Prolonged deparaffinization time and phenol/chloroform extraction did not influence DNA quality for PCR. Formalin fixed tissues can be successfully used for DNA/PCR of shorter fragments (318 bp) even if they are up to 39 years old. Longer fragment (720 bp) was successfully amplified from 1 and 10 years old specimens, also investigated whether DNA suitable for hybridization studies could be prepared from fixed tissue. Formalin caused irreversible DNA damages which were greater with prolonged fixation time making it unsuitable for hybridization studies, but still suitable for PCR amplification. Carnoy's and AMeX fixation resulted in consistently high yield of high molecular size DNA suitable for use in hybridization studies and PCR respectively. DNA from Papanicolaou stained smears was successfully amplified by PCR as well. Of all stains used in the preparation of smears, only eosin was detectable as a greenish band in agarose gels under ultraviolet illumination.

Brain Chemistry↗
PubMed1995

Nested polymerase chain reaction for detection of hepatitis C virus RNA in blood derivatives.

Testing for the presence of hepatitis C virus (HCV) in blood derivatives used in clinical medicine is of enormous importance to ensure the production of safe preparations. So far, no system has been developed for the isolation and detection of hepatitis C virus from blood derivatives. The recently introduced commercial kit for the detection of hepatitis C virus is designed for the isolation and detection of virus from blood serum. A reliable and reproducible method for the isolation of hepatitis C virus RNA, subsequent reverse transcription and nested polymerase chain reaction (PCR) from blood derivatives is described. Of 17 batches of factor VIII, gamma-globulin and anti tetanus, cytomegalovirus and Varicella-zoster immunoglobulin concentrates, respectively (14 negative for anti HCV and 3 of unknown anti HCV status) five were found positive in RNA-PCR.

Base Sequence↗
PubMed1994

p53 immunoreactivity in oligodendrogliomas.

Data are presented on p53 protein presence in human oligodendrogliomas whose progress from low grade to anaplastic oligodendroglioma can be followed. Expression was evaluated by formalin-fixed paraffin-embedded section immunohistochemistry, using monoclonal PAb 1801 antibody. The frequency of p53 protein accumulation is related to the stage of tumor malignancy. All the samples (100%) of malignant oligodendrogliomas were positive for p53 protein. Of 14 type II oligodendroglioma samples, 9 were positive (64%) while among type I oligodendroglioma the positivity was 28%. The mean proportion of reactive cells was also higher in malignant oligodendrogliomas. However, mean intensity staining did not differ among various grades of tumors. Our results point to the direct link between p53 protein accumulation and the malignant stage of human oligodendrogliomas. However, the value of p53 protein accumulation in predicting malignant behavior of oligodendrogliomas requires further confirmation.

Brain Neoplasms↗
PubMed1993

PCR amplification of DNA from stained cytological smears.

DNA from archival Papanicolaou stained smears was successfully amplified using the polymerase chain reaction (PCR) to see if it could be used for retrospective genome studies such as detection of the presence of human papilloma virus (HPV) and changes in p53 gene expression. DNA was isolated and purified by treatment with proteinase K, phenol/chloroform, and isoamyl alcohol. Segments of the human beta actin and TGF beta 1 gene were amplified by PCR. Of all stains used in the preparation of Papanicolaou smears, only eosin was detectable as a greenish band in ethidium bromide treated DNA gels under ultraviolet illumination.

Actins↗
PubMed1993

DNA amplification by polymerase chain reaction from brain tissues embedded in paraffin.

A method which enables analysis of DNA from archival paraffin embedded normal and malignant brain tissue is described. The demonstration of a 317-bp long beta-actin DNA sequence by the polymerase chain reaction (PCR) was used to identify which fixation procedure, deparaffinization time and DNA extraction procedure would give the best results. Tissue specimens 1-39 years old were included in the experiments. Specimens fixed in either 10% formalin, Carnoy's or AMeX fixative were found to be best suited for subsequent analysis by PCR. Paraformaldehyde and acetone compromised amplification efficiency, while Bouin's fixed tissue gave uniformly negative results. Regardless of fixative used, PCR reaction had to be run through at least 40 cycles. Prolonged deparaffinization time and phenol/chloroform extraction of DNA did not influence DNA quality as a template for PCR reaction. Formalin fixed brain tumours can be successfully used for DNA/PCR analysis even if they are up to 39 years old.

Base Sequence↗