CER 1 gene polymorphism in postmenopausal Roma and non-Roma Slovak women in connection with osteoporosis.

OBJECTIVE: The aim of our study was to investigate the relationship between the rs74434454 polymorphism of the CER1 gene and selected biochemical, densitometric and anthropometric markers in Slovak postmenopausal women of two ethnic groups: Roma and non-Roma. SUBJECTS AND METHODS: The scientific study included 303 postmenopausal women of the non-Roma and Roma populations who were divided into two groups based on densitometric measurements: control group (CG) and osteoporotic group (OG). Genomic DNA was isolated from peripheral blood using a commercial NucleoSpin® Blood kit following a standard protocol. The TaqMan Real-Time PCR method was used for genotyping. Biochemical markers were measured with Cobas e411 and Cobas Integra400 plus analysers. RESULTS: In the control group of postmenopausal Roma women, the occurrence of the risk genotype GG was not observed. In the group of Roma women with osteopenia and osteoporosis, the GG genotype occurred at a frequency of 3.03%. In the group of non-Roma women (between CG and OG) statistically significant differences were found in all monitored biochemical markers except CTx-I (p<0.66). In contrast, in the group of Roma women, statistical significance was only found in the osteoresorption marker CTx-I (p<0.007). In the population of Roma women, we did not find a statistically significant difference between the AA, AG and GG genotypes in any of the monitored markers. CONCLUSIONS: The results provide the first and unique insight on the distribution of genotypes and alleles of the rs74434454 CER1 gene polymorphism and its relationship to markers of bone metabolism in two ethnically distinct groups.

Preliminary results of ethnic divergence of G1181C (rs2073618) and C290T (rs9525641) OPG gene polymorphisms in groups of postmenopausal Slovak women.

OBJECTIVE: In this study, we focused on observation of the genetic polymorphisms of the OPG genes G1181C (rs2073618) and C290T (rs9525641), their interactions with biochemical markers and anthropometric parameters in groups of postmenopausal Slovak women (Roma and non-Roma, n = 311). PATIENTS AND METHODS: Genomic DNA was extracted and purified from peripheral blood leukocytes by the kit Ultraclean® Blood non-spin® (Carlsbad, CA, USA) using a standard protocol. Genotyping was performed by the TaqMan SNP genotyping assay. Biochemical markers were measured by the Cobas e411 (Roche Diagnostic, Tokyo, Japan) and Cobas Integra400 plus (Roche Diagnostic, Rotkreuz, Switzerland) analysers. RESULTS: We recorded a higher frequency of the T allele in the C290T polymorphism of the non-Roma control group (53.846%), in Roma groups: control (T - 56.618%) osteoporotic (T - 51.471%). In the G1181C polymorphism, the CC genotype occurred more in the osteoporotic group (34.286%) compared to the control group (27.885%). In the group of postmenopausal Roma women, a statistically significant difference (p<0.05) was found between osteoporotic and control in the biochemical parameters' osteocalcin, C-terminal telopeptide I, and age. Statistically significant differences (p<0.0001) were also found in bone mineral density and T-score. The high odds ratio suggests the association of G1181C with osteoporosis. A close relationship was found between haplotypes, BMD, T-score, and IL-6 in control; and BMI, WHR, T-score, and osteocalcin in osteoporotic groups of Roma and non-Roma women. CONCLUSIONS: The results point to differences in the occurrence of genotypes and associations of haplotypes with the manifestation of osteoporosis in Roma and non-Roma women. However, a larger number of samples is needed to determine whether or not there are differences between the Roma and non-Roma populations.

Subacute exposure to amygdalin influences compact bone remodeling of rabbits.

Amygdalin is most commonly occurring cyanogenic glycoside. It is found in seeds of many plant species. Our study was aimed to reveal whether pure intramuscularly injected amygdalin or apricot seeds peroral exposure cause changes in bone microstructure of rabbits. Twenty clinically healthy 5 months-old male rabbits were segregated into five groups. Animals from groups A1 and A2 were intramuscularly injected with amygdalin at doses of 0.6 and 3 mg/kg b.w. daily for 28 days. The groups S1 and S2 received commercial feed for rabbits mixed with crushed bitter apricot seeds at doses of 60 and 300 mg/kg b.w. during 28 days. The control (C) group did not receive any amygdalin. Intramuscular and peroral amygdalin administration did not affect total body weight, femoral length and femoral weight of rabbits. Similarly, microcomputed tomography (3D analysis) has shown that amygdalin had insignificant effect on relative bone volume, bone mineral density, cortical bone thickness, bone surface, trabecular thickness, trabecular number, trabecular separation. However, histological (2D analysis) revealed evident changes in compact bone microstructure of amygdalin-exposed rabbits consistent with a different vascularization and changed biomechanical properties. We can conclude that subacute exposure to amygdalin (both intramuscular and peroral) at the doses used in our study influenced compact bone remodeling.

Subacute exposure to alcohol in relation to bone microstructure of mice.

Our study aimed to investigate subacute exposure to alcohol in relation to bone microstructure of mice. Animals from experimental (E) group drank a solution composed of 15 % ethanol and water for 14 days (one remodeling cycle), while those from control (C) group drank only water. In the compact bone of E group, decreased bone formation and increased porosity were observed which corresponds with lower levels of serum alkaline phosphatase and glutathione. Alcohol significantly increased sizes of primary osteon's vascular canals and decreased those of secondary osteons, Haversian canals. Relative bone volume, bone mineral density (BMD), relative bone volume without marrow cavity were also lower in E group. On the contrary, trabecular bone microstructure did not differ significantly between E and C groups. Liver function test showed higher levels of alanine aminotransferase, aspartate aminotransferase in alcohol-fed mice. Serum calcium, phosphate were significantly lower in E group. According to our study, only changes in compact bone microstructure of mice following one remodeling cycle were observed due to both direct and indirect effects of alcohol.

Maternal relationship analysis of skeletal remains from Early Medieval cemetery in Slovakia.

SUMMARY: MtDNA from the skeletal remains of two bodies buried in the grave No. F44 from Nitra-Sindolka (woman and child) was analysed. Cemeteries in Sindolka belong to the Bijelo Brdo culture, where Slavs and Magyars were buried in the 10th - 11th centuries. Surprisingly, the analysis of mtDNA has shown no maternal relationship between buried persons, since the woman belongs to haplogroup T1a and the child to haplogroup J. Since both haplogroups can be found in Near Eastern and European populations at similar frequencies, the tested samples from the tomb No. F44 cannot be clearly assigned to ethnic origin.

Femoral bone microstructure in 1-month-old non-transgenic versus transgenic rabbits with the WAP-hFVIII gene construct.

Differences in microscopic structure of the femur between 1-month-old transgenic rabbits carrying the hFVIII gene and non-transgenic rabbits were investigated. Bone microstructure was evaluated from the point of view of qualitative and quantitative histological characteristics. We identified fibrolamellar bone tissue only in the transgenic animals. Measured values for area, perimeter of the Haversian canals and minimum diameter of the primary osteons' vascular canals were higher in 1-month-old transgenic individuals (P < 0.05; P < 0.001). We also observed lower concentrations of Ca, P, K, solids, and total mineral content in femora of transgenic rabbits. A statistically significant difference was observed for the concentration of Ca (P < 0.05). Our results indicate evident changes in both qualitative and quantitative histological characteristics of the femur, which result especially in better blood supply and slightly reduced mineralization process in 1-month-old transgenic rabbits.

Comparative study of compact bone tissue microstructure between non-transgenic and transgenic rabbits with WAP-hFVIII gene construct.

The aim of this study was to investigate differences in compact bone tissue microstructure between New Zealand White transgenic rabbits with human blood clotting factor VIII gene and the non-transgenic ones. The transgene was under the transcriptional control of the whey acidic protein (WAP) promoter. Altogether, 42 femora were analysed. Specimens were prepared using standard histological equipment, producing thin sections approximately 80-100 microm. Areas, perimeters, minimum and maximum diameters of vascular canals of primary osteons, Haversian canals and secondary osteons were measured. Our results indicate the basic structural pattern of the bone tissue was primary vascular longitudinal in both groups of rabbits. However, a fibrolamellar bone tissue was identified only in the transgenic rabbits. The measured variables of the vascular canals of the primary osteons, the Haversian canals and the secondary osteons were higher in transgenic rabbits in comparison with non-transgenic ones in most cases and the differences were often statistically significant (P < 0.01; P < 0.001). More significant differences were noticed in males (transgenic versus non-transgenic). However, it is not easy to find the real cause of the changes. Despite the fact that no hFVIII mRNA expression was found in the analysed bone of transgenic rabbits, we presume that the observed differences could be associated with transgenesis. In this article, we discuss possible associations between transgene integration as well as aneuploidy and/or observed microstructural changes in compact bone tissue.

Changes of femoral bone tissue microstructure in transgenic rabbits.

Bone tissue microstructure of femur was investigated in transgenic New Zealand White rabbits with human factor VIII gene. Altogether 42 bones (24 from transgenic rabbits and 18 from non-transgenic ones) were analysed. Specimens were prepared using standard histological equipment, producing thin sections of approximately 80-100 microm. For histomorphometrical analysis areas, perimeters, minimum and maximum diameters of osteons' vascular canals and of osteons were measured. We found out that the basic structural pattern of femoral bone tissue was primary vascular longitudinal in both groups of rabbits. However, a new type of the bone tissue--fibrolamellar--was identified only in the transgenic rabbits. The measured variables of the osteons' vascular canals were higher in transgenic individuals in comparison with the nontransgenic ones (except for maximal diameter) and the differences were statistically significant (P < 0.05; P < 0.01). We suppose that the observed differences could be associated with transgenesis. In an effort to explain these differences we compared the cytogenetic profile of bone marrow cells between transgenic and non-transgenic rabbits. A significantly higher rate of aneuploidy was observed in c-metaphase spreads of transgenic individuals as compared to non-transgenic ones (P < 0.001). Despite the fact that no hFVIII mRNA expression was found in the femur of transgenic rabbits, we discussed an association of transgene integration into the genome and microstructural changes in the bone. In any case, the results indicate that transgenesis can also produce changes in other tissues than in the target ones.

A simple and inexpensive DNA purification for malignant hyperthermia PCR detection in porcine hair roots.

We isolated DNA from porcine hair roots by silica matrix. Purified DNA is suitable for PCR analysis of malignant hyperthermia in the pig breeds. The using of hair roots as a source of DNA provides noninvasivity and it is also convenient for breeders. We compared amplification of DNA purified by silica matrix and Prep-A-gene kit. DNA purification by silica is as effective as Prep-A-gene purification and it is advantageous because of the low price.

Meat quality assessed Ante- and Post mortem by different ryanodine receptor gene status of pigs.

Different breeds and crossbreeds (total 63) of pigs with defined genotype (DNA based test on RYR 1 gene status) were divided in normal (non mutant), heterozygotes (carriers, monomutant) and homozygote positive (dimutant) animals. Muscle biopsy samples were taken at approximately 80 kg live weight using an efficient non-stress spring-loaded biopsy instrument. After incubation of muscle samples (39 °C, 1 hr) analyses pH, WHC (fluid), R value (ITP:ATP) were performed. After slaughter, meat quality values (pH(1), pH(24), R value, colour, drip loss) were also carried out. Heterozygotes were intermediate and between both homozygote pig groups in most meat quality values analysed. Using WHC (fluid volume) values (0.45 and 0.48 respectively) it was possible to divide heterozygotes into two groups, each closer to homozygotes. The values of pH, WHC (fluid), as measured at definite times on the biopsies, were correlated with the pH(1) post-mortem value (r approx 0.6-0.7). The results support previous findings on the possibility of differentiating pigs within DNA test-defined groups. Introducing the gene into a pig population could be useful, in improving leanness and muscling: but meat quality is certain to deteriorate. Further experiments are needed to support the possibility of identifying PSE-prone pigs from among normal non mutant pigs.