New reference values must be established for the Alberta Infant Motor Scales for accurate identification of infants at risk for motor developmental delay in Flanders.

BACKGROUND: The Alberta Infant Motor Scales (AIMS) is a reliable and valid assessment tool to evaluate the motor performance from birth to independent walking. This study aimed to determine whether the Canadian reference values on the AIMS from 1990-1992 are still useful tor Flemish infants, assessed in 2007-2010. Additionally, the association between motor performance and sleep and play positioning will be determined. METHODS: A total of 270 Flemish infants between 0 and 18 months, recruited by formal day care services, were assessed with the AIMS by four trained physiotherapists. Information about sleep and play positioning was collected by mean of a questionnaire. RESULTS: Flemish infants perform significantly lower on the AIMS compared with the reference values (P < 0.001). Especially, infants from the age groups of 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 and of 15 months showed significantly lower scores. From the information collected by parental questionnaires, the lower motor scores seem to be related to the sleep position, the amount of play time in prone, in supine and in a sitting device. Infants who are exposed often to frequently to prone while awake showed a significant higher motor performance than infants who are exposed less to prone (<6 m: P = 0.002; >6 m: P = 0.013). Infants who are placed often to frequently in a sitting device in the first 6 months of life (P = 0.010) and in supine after 6 months (P = 0.001) performed significantly lower than those who are placed less in it. CONCLUSION: Flemish infants recruited by formal day care services, show significantly lower motor scores than the Canadian norm population. New reference values should be established for the AIMS for accurate identification of infants at risk. Prevention of sudden infant death syndrome by promoting supine sleep position should go together with promotion of tummy time when awake and avoiding to spent too much time in sitting devices when awake.

Test-retest reliability of the assessment of postural stability in typically developing children and in hearing impaired children.

The purpose of this study was to establish test-retest reliability of centre of pressure (COP) measurements obtained by an AccuGait portable forceplate (ACG), mean COG sway velocity measured by a Basic Balance Master (BBM) and clinical balance tests in children with and without balance difficulties. 49 typically developing children and 23 hearing impaired children, with a higher risk for stability problems, between 6 and 12 years of age participated. Each child performed the modified Clinical Test of Sensory Interaction on Balance (mCTSIB), Unilateral Stance (US) and Tandem Stance on ACG, mCTSIB and US on BBM and clinical balance tests: one-leg standing, balance beam walking and one-leg hopping. All subjects completed 2 test sessions on 2 different days in the same week assessed by the same examiner. Among COP measurements obtained by the ACG, mean sway velocity was the most reliable parameter with all ICCs higher than 0.72. The standard deviation (SD) of sway velocity, sway area, SD of anterior-posterior and SD of medio-lateral COP data showed moderate to excellent reliability with ICCs between 0.55 and 0.96 but some caution must be taken into account in some conditions. BBM is less reliable but clinical balance tests are as reliable as ACG. Hearing impaired children exhibited better relative reliability (ICC) and comparable absolute reliability (SEM) for most balance parameters compared to typically developing children. Reliable information regarding postural stability of typically developing children and hearing impaired children may be obtained utilizing COP measurements generated by an AccuGait system and clinical balance tests.

The Smc5/6 complex: more than repair?

Through its functions in chromosome replication, segregation, and repair, the Smc5/6 complex has a central role in the maintenance of genome stability. The complex is part of the family of structural maintenance of chromosome protein complexes that also includes cohesin and condensin. Mutations in any of these complexes disrupt chromosome segregation and render cells hypersensitive to different types of DNA damage. The chromosome mis-segregation phenotypes in cohesin and condensin mutants can be attributed to their functions in sister chromatid cohesion and chromosome condensation, respectively. Cohesin-dependent chromatid cohesion is also needed for DNA double-strand break repair, whereas condensin is required for repair of single-strand breaks. How Smc5/6 promotes chromosome stability is largely unknown. Accumulating data suggest that it prevents accumulation of aberrant DNA links between sister chromatids created during repair by homologous recombination. A long-standing idea is that it also has a role in the maintenance of nondamaged chromosomes. Here, we present an overview of the current knowledge of Smc5/6 and discuss a possible nonrepair role of the complex.

Nej1p, a cell type-specific regulator of nonhomologous end joining in yeast.

Mutant yeast strains lacking the silencing proteins Sir2p, Sir3p, or Sir4p have a defect in a DNA double-strand break (DSB) repair pathway, called nonhomologous end joining (NHEJ). Mutations in sir genes also lead to the simultaneous expression of a and alpha mating type information, thus generating a nonmating haploid cell type with many properties shared with a/alpha diploids. We addressed whether cell type or Sir proteins per se regulate NHEJ by investigating the role of a novel haploid-specific gene in NHEJ. This gene, NEJ1, was required for efficient NHEJ, and transcription of NEJ1 was completely repressed in a/alpha diploid and sir haploid strains. The NEJ1 promoter contained a consensus binding site for the a1/alpha2 repressor, explaining the cell type-specific expression. Expression of Nej1p from a constitutive promoter in a/alpha diploid and sir mutant strains completely rescued the defect in NHEJ, thus showing that Sir proteins per se were dispensable for NHEJ. Nej1p and Lif1(P), the yeast XRCC4 homolog, interacted in two independent assays, and Nej1p localized to the nucleus, suggesting that Nej1p may have a direct role in NHEJ.

Kluyveromyces lactis Sir2p regulates cation sensitivity and maintains a specialized chromatin structure at the cryptic alpha-locus.

In Saccharomyces cerevisiae, transcriptional silencing of the cryptic mating type loci requires the formation of a heterochromatin-like structure, which is dependent on silent information regulator (Sir) proteins and DNA sequences, called silencers. To learn more about silencing, we characterized the mating type loci from the yeast Kluyveromyces lactis. The K. lactis MAT, HMRa, and HMLalpha loci shared flanking DNA sequences on both sides of the loci presumably acting as recombinational targets during mating type switching. HMRa contained two genes, the a1 gene similar to the Saccharomyces a1 gene and the a2 gene similar to mating type genes from other yeasts. K. lactis HMLalpha contained three genes, the alpha1 and alpha2 genes, which were similar to their Saccharomyces counterparts, and a novel third gene, alpha3. A dam-methylase assay showed Sir-dependent, but transcription-independent changes of the chromatin structure of the HMLalpha locus. The HMLalpha3 gene did not appear to be part of the silent domain because alpha3p was expressed from both MATalpha3 and HMLalpha3 and sir mutations failed to change the chromatin structure of the HMLalpha3 gene. Furthermore, a 102-bp silencer element was isolated from the HMLalpha flanking DNA. HMLalpha was also flanked by an autonomously replicating sequence (ARS) activity, but the ARS activity did not appear to be required for silencer function. K. lactis sir2 strains grown in the presence of ethidium bromide (EtBr) accumulated the drug, which interfered with the essential mitochondrial genome. Mutations that bypassed the requirement for the mitochondrial genome also bypassed the EtBr sensitivity of sir2 strains. Sir2p localized to the nucleus, indicating that the role of Sir2p to hinder EtBr accumulation was an indirect regulatory effect. Sir2p was also required for growth in the presence of high concentrations of Ni(2+) and Cu(2+).

Pharmacodynamics of cisplatin in human head and neck cancer: correlation between platinum content, DNA adduct levels and drug sensitivity in vitro and in vivo.

Total platinum contents and cisplatin-DNA adduct levels were determined in vivo in xenografted tumour tissues in mice and in vitro in cultured tumour cells of head and neck squamous cell carcinoma (HNSCC), and correlated with sensitivity to cisplatin. In vivo, a panel of five HNSCC tumour lines growing as xenografts in nude mice was used. In vitro, the panel consisted of five HNSCC cell lines, of which four had an in vivo equivalent. Sensitivity to cisplatin varied three- to sevenfold among cell lines and tumours respectively. However, the ranking of the sensitivities of the tumour lines (in vivo), also after reinjection of the cultured tumour cells, did not coincide with that of the corresponding cell lines, which showed that cell culture systems are not representative for the in vivo situation. Both in vitro and in vivo, however, significant correlations were found between total platinum levels, measured by atomic absorption spectrophotometry (AAS), and tumour response to cisplatin therapy at all time points tested. The levels of the two major cisplatin-DNA adduct types were determined by a recently developed and improved 32P post-labelling assay at various time points after cisplatin treatment. Evidence is presented that the platinum-AG adduct, in which platinum is bound to guanine and an adjacent adenine, may be the cytotoxic lesion because a significant correlation was found between the platinum-AG levels and the sensitivities in our panel of HNSCC, in vitro as well as in vivo. This correlation with the platinum-AG levels was established at 1 h (in vitro) and 3 h (in vivo) after the start of the cisplatin treatment, which emphasizes the importance of early sampling.

The potential of plantinum-DNA adduct determination in ex vivo treated tumor fragments for the prediction of sensitivity to cisplatin chemotherapy.

BACKGROUND: Response to cisplatin-therapy is assumed to be related to the formation of platinum (Pt)-DNA adducts. Measurement of these adducts prior to therapy could be of value to improve cisplatin based cancer therapy. MATERIALS AND METHODS: We determined Pt-GG and Pt-AG adduct levels by use of 32P-postlabeling after ex vivo cisplatin treatment of fragments of head and neck squamous cell carcinoma (HNSCC) xenografts (five lines), and of tumor biopsies from patients with HNSCC (n = 8) and testicular cancer (n = 8). RESULTS: Adduct levels in fragments (3 x 3 x 3 mm) exposed to 10 to 80 microM cisplatin for one hour, showed positive correlations with the in vivo response to cisplatin treatment (P < 0.05), as well as with the xenograft adduct levels observed after in vivo cisplatin treatment (P < 0.02). After an additional five-hour drug-free incubation period the correlations were absent. When patient tumor fragments were exposed ex vivo to 80 microM cisplatin for one hour, adduct levels were similar in HNSCC and testicular cancer. Persistence of adducts was observed for testicular cancer in the additional drug-free period. The adduct levels in the samples of two HNSCC patients who received cisplatin chemotherapy were in line with the hypothesis that higher adduct levels are associated with a better response. CONCLUSION: Our preliminary results show that analysis of DNA adducts following ex vivo drug treatment is a feasible approach towards a predictive assay, which warrants further investigation.

The growth inhibiting effect of docetaxel (Taxotere) in head and neck squamous cell carcinoma xenografts.

Docetaxel (Taxotere) is a new antitumor agent with activity in patients with ovarian, lung and breast cancer. In this study, docetaxel was tested for its antitumor effect in human tumor xenografts derived from head and neck squamous cell carcinoma (HNSCC). A significant growth inhibiting effect was observed in the two tested lines, at the well tolerated dose of 20 m/kg docetaxel. Two intravenous injections were given with a week interval. Both lines were less sensitive to treatment with cisplatin, indicating that no cross-reactivity exists between these drugs. Docetaxel has a promising outlook in the treatment of patients with head and neck squamous cell carcinoma (HNSCC).

Importance of pharmacodynamics in the in vitro antiproliferative activity of the antifolates methotrexate and 10-ethyl-10-deazaaminopterin against human head and neck squamous cell carcinoma.

The pharmacodynamic profiles of methotrexate (MTX) and 10-ethyl-10-deazaaminopterin (10-EdAM) were determined in three head and neck squamous cell carcinoma (HNSCC) cell lines. Cell growth inhibition was tested using a semi-automated 96-well based proliferation assay, the sulforhodamine B (SRB) assay. Drug concentrations ranged from 10(-5) to 10(-9) M, with exposure periods of 4, 24, 48, 72 and 96 hr. The SRB-test was performed after each of these periods of continuous exposure and after an additional period of 24 and 48 hr in drug-free medium. Without a drug-free period the IC50 values strongly depended on the time of exposure. For example, with respect to MTX, IC50 values at 24 hr ranged from 2.9 (UM-SCC-14C) to over 10 microM (UM-SCC-22B and -11B), but when exposed continuously for 96 hr, IC50 values varied between 0.039 and 0.1 microM. 10-EdAM followed a similar sensitivity pattern with 5-20-fold lower IC50 values. The minimal time to achieve significant growth inhibition varied between the cell lines, < 24 hr for UM-SCC-14C, > 24 and > 48 hr for UM-SCC-11B and -22B, respectively. The cell lines also varied with respect to growth behaviour when placed in drug-free medium for an additional period. Growth of UM-SCC-14C cells was recovered significantly after removing the drug, whereas UM-SCC-22B showed a different pattern: when cultured for over 48 hr, cell growth was strongly inhibited, independent of the drug being removed. This variable pattern of sensitivity could be correlated with the capacity of the cells to form polyglutamate derivatives. After 24 hr, drug accumulation was at least three times lower in UM-SCC-14C than in both other cell lines. The low level of antifolate accumulation in UM-SCC-14C is in line with the recovery from growth inhibition at culture in drug-free medium, while the persistent growth inhibition observed in UM-SCC-22B agrees with the intracellular accumulation of higher polyglutamates. In conclusion, these experiments show that the pharmacodynamic profile varies between HNSCC cell lines and plays an important role in the growth inhibition by antifolates. Both exposure time and the intrinsic capacity to synthesize polyglutamates are important factors in the sensitivity of HNSCC to antifolate drugs.