Flow cytometric DNA hypertetraploidy tends to be more frequent in male than in female breast cancers.

The aim of the study was to explore possible differences in DNA flow cytometric characteristics, particularly differences in distribution of DNA indices of aneuploid clones, between male and female breast cancers. We retrospectively analyzed 31 male breast cancers. Clinicopathological and DNA flow cytometric characteristics of male breast cancers (patient age, tumor size, histological type, histological grade, axillary lymph node status, hormone receptor expression, ploidy, and S-phase fraction) were compared with that of the control group of matched female breast cancers. Hormone receptors and HER-2/neu were investigated immunohistochemically with additional chromogenic in situ hybridization (CISH) analysis of HER-2/neu 2+ cases. Ploidy and S-phase fraction were determined by DNA flow cytometry. Comparison with clinicopathological features was made using χ (2) and t test. Aneuploidy was found in 78% of the cases, with the predomination of hypotetraploid clones (39%), followed by tetraploid (23%) and hypertetraploid clones (16%). We found higher frequency of hypertetraploidy in male breast cancers (16 and 6%, respectively) than in the control group of matched female breast cancers. Clinicopathological features of hypertetraploid male breast cancers did not differ from that of non-hypertetraploid cancers. Higher frequency of hypertetraploidy among male breast cancers might indicate different cytogenetical evolutionary pathway between male and female breast cancer.

Skeletal remains from late Roman period: "as old as Diocletian's Palace".

In 2000, human skeletal remains were discovered in Split (Croatia). As archaeologists confirmed, it was an ancient skeleton accompanied by ceramics and bracelet characteristic for late Roman period whose possible violent death was excluded. The bone sample was radiocarbon dated by AMS to 1750 years. DNA was successfully extracted from the bone sample and subsequently typed using mt DNA and STR systems. The metal content was determined by atomic absorption spectrometry (AAS) in flame mode. Mercury concentration was determined by direct consecutive measures taken with a mercury analyzer. According to our results, we consider that the bones could belong to the one of the last citizens of the Diocletian's Palace.

Near-diploid hyperploidy in early breast cancer (T1a,b) is associated with higher risk of lymph node involvement.

Due to the worldwide implementation of the mammographic screening program early breast cancer (T1a,b) has become more prevalent form of breast cancer. Although T1a,b breast cancers are generally associated with excellent prognosis, some of them, particularly those with lymph node involvement, has unfavourable outcome. Searching for additional prognostic factors, we investigated DNA content of 163 T1a,b cancers measured by DNA flow cytometry, and correlated it with regional lymph node status. T1a,b cancers were divided into four ploidy classes based on their DNA index (DI): hypodiploid (DI < 0.95), diploid (DI 0.95-1.05), low-hyperploid (DI 1.06-1.3), and high-hyperploid (DI > 1.3). Diploid T1a,b cancers were associated with negative lymph node status (p = 0.003). Among aneuploid cancers only low-hyperploid tumors were associated with positive lymph node status (p = 0.03). The histopathological features of low-hyperploid group of T1a,b cancers did not differ from the other three ploidy groups of cancers, except for lower S-phase fraction of tumor cells in low-hyperploid group compared to high-hyperploid group (p = 0.01). Our data showed that near-diploid hyperploid T1a,b cancers are associated with higher risk of lymph node involvement despite similar clinicopathological features shared with other ploidy classes of T1a,b tumors.

Can we identify the group of small invasive (T1a,b) breast cancers with minimal risk of axillary lymph node involvement? A pathohistological and DNA flow cytometric study.

The goal of this study was to identify a group of small (≤1cm) breast cancers (T1a,b) with a particularly low probability of axillary lymph node metastases, where routine axillary staging may be unnecessary. We retrospectively analyzed 152 T1a,b breast carcinomas with axillary dissection surgically removed at Clinical Hospital Center Split (Croatia) in the period from 1997 to 2006. The analysis included 40 T1a,b cancers with, and 112 T1a,b cancers without axillary lymph node metastases. The basic morphological features of cancers were investigated histologically, while hormone receptors and HER2/neu were investigated immunohistochemically with an additional CISH analysis of HER2/neu 2+ cases. The ploidy and S-phase fraction were determined by DNA flow cytometry. The association of the investigated features with the likelihood of axillary lymph node metastases was analyzed by univariate and multivariate analysis. The univariate analysis showed that lymph node metastases were associated with tumor size (T1a/T1b; p=0.026), histological type (ductal/non-ductal; p=0.014), lymphovascular invasion (p<0.001), HER2/neu expression (p=0.04), ploidy (p=0.027), and combined values of ploidy and S-phase fraction (p=0.025). The lymphovascular invasion was the only independent factor associated with axillary nodal metastases (p=0.01). In the group of T1a,b cancers without lymphovascular invasion, HER2/neu expression (p=0.021) and combined values of ploidy and S-phase fraction (p=0.016) were independent factors associated with axillary lymph node metastases. This study showed that diploid T1a,b cancers with low S-phase fraction, which are also without lymphovascular invasion and HER2/neu amplification, represented the group of cancers with a low probability of axillary lymph node metastases.

Discovering the 60 years old secret: identification of the World War II mass grave victims from the island of Daksa near Dubrovnik, Croatia.

AIM: To describe the organization, field work, forensic anthropological examination, and DNA analysis conducted to identify the victims from a World War II mass grave found on the Dalmatian island of Daksa near Dubrovnik (Croatia) in 2009. METHODS: Excavation of the site was performed according to standard archeological procedures. Basic anthropological examination was made to determine the minimum number of victims, sex, age at death, and height. The bones with pathological and traumatic changes were identified. DNA was extracted from powdered bones and relatives' blood samples. Y-chromosome and autosomal short tandem repeats (STR) were used to establish the relationship of the remains with the putative family members. RESULTS: The remains were found to belong to at least 53 distinctive victims. All were male, mostly with gunshot wounds to the head. DNA analysis and cross-matching of the samples with relatives resulted in 14 positive identifications using the Y-chromosomal STRs and 4 positive identifications using the autosomal STRs. CONCLUSIONS: This study showed that even in cases of more than 50-year-old, highly degraded human remains from mass graves, Y-chromosomal and autosomal STRs analysis can contribute to identification of the victims.

Y-chromosomal short tandem repeat haplotypes in southern Croatian male population defined by 17 loci.

AIM: To define the Y-chromosome genetic structure in a sample of men from southern Croatia. METHODS: Blood samples were collected from 166 unrelated healthy men from southern Croatia at the Department of Forensic Medicine and Biochemical Laboratory of University Hospital Split between 2004 and 2007. Genomic DNA was extracted using the standard procedures. Seventeen Y-chromosome short tandem repeat (Y-STR) polymorphic loci (DYS456, DYS389I, DYS390, DYS389II, DYS458, DYS19, DYS385, DYS393, DYS391, DYS439, DYS635, DYS392, GATAH4, DYS437, DYS438, and DYS448) were analyzed using AmpFlSTR Yfiler Polymerase Chain Reaction Amplification Kit. RESULTS: We observed 152 different haplotypes. Total haplotype diversity was 0.997289 and 141 haplotypes (84.49%) were unique. The most common haplotype was shared by only 4 men in the study sample. The locus diversity ranged between 0.21292 for DYS392 and 0.75546 for DYS439 locus. CONCLUSION: The Y-chromosome structure in men from southern Croatia is very diverse. Combination of Y chromosome 17 STR loci may be used as a powerful tool for individual identification and parentage analysis in the southern Croatian male population.