CASE-REPORT Association between an ACAN gene variable number tandem repeat polymorphism and lumbar disc herniation: a case control study.

We investigated the association between an aggrecan gene (ACAN) polymorphism and lumbar disc herniation (LDH). This was a case-control study with quinquennial age and gender groups. The study comprised 119 men and women aged between 20 and 60 from Goiânia (Brazil). Of these, 39 were allocated to the case group (Ca) and 80 to the control group (Ct). We gathered sociodemographic and clinical data, and peripheral blood samples. DNA was isolated for genotyping the ACAN variable number tandem repeat (VNTR) via conventional polymerase chain reaction (PCR). Data were statistically analyzed using the chi-square test, multiple comparison analysis, the Student t-test, and odds ratios, with a level of significance set at 5% (P ≤ 0.05). The groups were homogenous in terms of sociodemographic, anthropometric, and life style variables. The allele score for the ACAN VNTR was significantly lower in volunteers with LDH; the A22 allele was significantly more prevalent in this same group; the Ca group presented greater frequency of short alleles A13-A25, whereas the Ct group presented a higher frequency of long alleles. However, this difference was not statistically significant. In both groups, the most common alleles were A28, A27, and A29, and the A26/A26 genotype was significantly more common in the Ca group. The results showed an association between short alleles and LDH among the investigated adults (Ca), corroborating the hypothesis that aggrecan with shorter repeat lengths can lead to a reduction in the physiological proteoglycan function of intervertebral disc hydration and, consequently, increased individual susceptibility to LDH.

Effect of cell cycle position on dexamethasone binding by mouse and human lymphoid cell lines: correlation between an increase in dexamethasone binding during S phase and dexamethasone sensitivity.

We determined the effect of cell cycle position on the amount of dexamethasone that was specifically bound by mouse and human lymphoid cell lines. Cell lines that were either sensitive or resistant to growth inhibition by dexamethasone were compared. Exponentially growing cells were separated by centrifugal elutriation into fractions that corresponded to different positions in the cell cycle. The cell cycle phase distribution of each fraction was estimated by flow cytometry and autoradiography. The amount of dexamethasone bound per cell in each fraction was measured by a whole cell binding assay. In three dexamethasone-sensitive cell lines (two mouse and one human), we found that the amount of dexamethasone bound per cell increased 2-4-fold between G1 phase and S phase, and then decreased during G2/M phase. Results were the same when the amount of dexamethasone bound per milligram of cell protein was measured. Binding affinity was the same during G1 phase and S phase, but the proportion of bound dexamethasone that translocated to the nucleus was greater during S phase. In contrast, we found that the amount of dexamethasone bound per cell by three dexamethasone-resistant cell lines (two mouse and one human) did not increase during S phase. Our results indicate that cell cycle changes in dexamethasone binding are not simply related to changes in cell protein or cell volume during the cell cycle. An increase in dexamethasone binding during S phase may be required for dexamethasone to inhibit cell growth, and a failure of dexamethasone binding to increase during S phase might represent a new mechanism of dexamethasone resistance in lymphoid cells.

Centrifugal elutriation of human lymphoid cells: synchronization and separation of aneuploid cells into diploid and tetraploid populations.

Both normal and leukemic human lymphoid cell lines were separated into populations corresponding to different positions in the cell cycle by centrifugal elutriation. Each population was analyzed for cell concentration, cell volume, [3H]thymidine incorporation, percentage S phase by autoradiography, and percent G1, S, and G2/M phases by flow cytometry. The smallest cells, collected at the lowest flow rate, were in G1 phase. Cells collected at increasing flow rates progressively increased in volume and represented distinct positions in the cell cycle transition from G1 phase, through S phase, and into G2/M phase. The purity of the G1 population varied according to cell load. One hundred percent of cells were recovered and cells collected in G1- and S-phase populations proliferated in culture with patterns characteristic of synchronized cells. An aneuploidy leukemia cell line, CEM, was separated into near-diploid and near-tetraploid populations by centrifugal elutriation. This method of cell separation provides large numbers of human lymphoid cells at different positions in the cell cycle for investigating the relationship between the cell cycle and various surface membrane and metabolic properties of cells. Aneuploid leukemia and lymphoma cells can be separated by centrifugal elutriation into populations which contain different numbers of chromosomes for comparisons of their biologic properties.