Expression of γ-H2AX and patient prognosis in breast cancer cohort

H2AX phosphorylation is a novel marker of DNA double-stranded breaks. In the present study, we assessed the γ-H2AX expression, its association with other clinicopathologic characteristics, and the prognosis in a cohort of 97 patients with breast cancer. Ninety-seven specimens of tumor tissue and 77 adjacent normal tissues from patients with breast cancer were examined. All patients underwent modified radical mastectomy or local tumor resection without lymph node dissection. γ-H2AX expression was assessed by standard immunohistochemistry. Patients were followed after surgery for a mean duration of 70.1 ± 18.7 months (range, 6-93 months). The γ-H2AX staining was positive in 27 (27.8%) patients. The positive rates of H2AX were 26.0% and 2.6% in tumor tissue and adjacent normal tissues, respectively. γ-H2AX positive status was negatively associated with TNM staging, with 24 positive cases (32.4%) in TNM staging I-II, while no positive cases in TNM staging III-IV (P = 0.026). Sixteen patients (16.5%) died during the follow-up. No significant association between γ-H2AX expression and patient survival was detected. The unadjusted HR (hazard ratio) for γ-H2AX positive was 0.84 (95% CI: 0.27, 2.60). In TNM staging subgroup analysis, death only occurred in γ-H2AX negative patients. Our study is the first study to demonstrate that expression of γ-H2AX is associated with TNM staging. Due to the small sample and limited follow-up time, we did not observe a significant association between γ-H2AX and patient survival. γ-H2AX expression could be a potential biomarker for cancer diagnosis and prediction, and further studies are in need.     

Developing dendritic cell polynucleotide vaccination for prostate cancer immunotherapy.

Immunotherapy has been successfully used to treat some human malignancies, principally melanoma and renal cell carcinoma. Genetic-based cancer immunotherapies were proposed which prime T lymphocyte recognition of unique neo-antigens arising from specific mutations. Genetic immunization (polynucleotide vaccination, DNA vaccines) is a process whereby gene therapy methods are used to create vaccines and immunotherapies. Recent findings indicate that genetic immunization works indirectly via a bone marrow derived cell, probably a type of dendritic antigen presenting cell (APC). Direct targeting of genetic vaccines to these cells may provide an efficient method for stimulating cellular and humoral immune responses to infectious agents and tumor antigens. Initial studies have provided monocytic-derived dendritic cell (DC) isolation and culture techniques, simple methods for delivering genes into these cells, and have also uncovered potential obstacles to effective cancer immunotherapy which may restrict the utility of this paradigm to a subset of patients.     

Isolation and culture of protoplasts from cotyledons of Pinus coulteri D. Don.

A protocol was developed for the isolation and culture of protoplasts from the cotyledons of seedlings of Pinus coulteri D. Don. Incubation of cotyledon pieces in a mixture consisting of cellulase Onozuka R10 2%, Pectolyase Y-23 0.1%, mannitol 10%, CaCl₂ 500 mg/l and other macro and micro-nutrients yielded viable protoplasts. After 24 hours of culture in a complex nutrient medium, the protoplasts regenerated new cell walls and the first divisions were observed within 7–10 days. Small cell colonies were formed within 15–20 days, but these started to accumulate phenolics and no further growth of the colonies was observed.     

Cytochemical Assay for Differential Respiratory Activity in Roots and Root Hairs

The study of the underground parts of plants is often difficult, and as a result roots are often treated as homogeneous physiological entities with respect to root respiration. In this study we demonstrate a partitioning of respiration within root tissues using nitro blue tetrazo-lium staining and an incident light optical system that permits detailed observations of intact roots. The assay is rapid and easy to perform, and reveals that respiratory activity in roots is not uniform in space and time. The results show that root hairs in particular may be regions of enhanced respiratory activity in some species or in certain developmental or physiological states. This fact has important implications for the role of root hairs in the overall respiratory budget of roots and the energetics of nutrient assimilation. The results suggest that root respiration studies should consider differential respiratory activities of root cell types within roots.     

Photosynthetic characteristics of photoautotrophically grown tobacco callus cells

Haploid callus cells of tobacco (Nicotiana tabacum) were grown photoautotrophically on a solid agar medium in the absence of sucrose in Petri plates in an atmosphere of 1% or 3% CO₂ in air. The averages of dry weight increases for four to five consecutive passages were 2.3- to 3.6-fold per 3-week passage for different subclones. Photosynthetic ¹⁴CO₂ assimilation was maximum at about 1% CO₂ with half-maximal rates obtained at 0.2% CO₂. At saturating CO₂ concentration the average rate of CO₂ fixation was about 5 μmole per gram fresh weight per hour or about 125 μmole per mg of chlorophyll per hour.     

Isolation and Characterization of Glycine Hydroxamate-resistant Cell Lines of Nicotiana tabacum

Seven lines of haploid Nicotiana tabacum tissue culture selected for resistance to normally toxic levels of the glycine analog glycine hydroxamate, a competitive inhibitor of the glycine decarboxylase reaction, were investigated. The presence of glycine hydroxamate greatly increased the intracellular concentration of both glycine and alanine in wild type and resistant cell lines, suggesting that the inhibitor blocks both glycine- and alanine-utilizing reactions. All the resistant cell lines, whether grown in the presence or absence of glycine hydroxamate, had high intracellular concentrations of the 12 free amino acids which were analyzed, including glycine and serine. (These lines averaged 3.6 times the total amino acid content of wild-type cells in the absence of the inhibitor). The resistant cell lines were indistinguishable from wild-type cell lines in their metabolism of radioactively labeled glycine hydroxamate and glycine. Comparison of the metabolism of radioactively labeled alanine, glycolate, and glyoxylate in wild-type and α resistant line also revealed no distinctive differences. Glycine decarboxylase activities were unaltered in the resistant cell lines. The cellular toxicity of glycine hydroxamate is considered in relation to (1) the competitive inhibition by glycine hydroxamate of the glycine- and alanine-utilizing enzymes and (2) the resultant imbalances caused by high intracellular concentrations of these amino acids. The significance of elevation of total free amino acid concentration in effecting resistance to the inhibitor is discussed.     

Posters

Introduction  Fine needle aspiration (FNA) cytology of the thyroid is a well-established test in the clinical work-up of patients with solitary nodules of the thyroid. Thyroid FNA does however have limitations and audit of diagnostic performance is important.
Methods  The histopathology archives of the Royal Victoria Hospital were searched for all thyroid resections and the histopathological diagnosis was correlated with the pre-operative cytological diagnosis, where available. Special emphasis was placed on the accuracy of tumour diagnosis.
Results  A total of 173 cases were identified during the 2-year period, of these 93 had available pre-operative FNA. A total of 57 tumours were identified. A small number (six of 57) of significant discrepancies were identified. These included a malignant lymphoma diagnosed as Hashimoto's thyroiditis, a metastasis which the FNA had suggested was a medullary carcinoma and an insular carcinoma diagnosed as medullary carcinoma on FNA. False positives included a colloid cyst diagnosed as suspicious of malignancy and a cytological diagnosis of papillary carcinoma not confirmed on histology.
Discussion  At present, the majority of thyroid FNAs in our clinics are performed by surgeons and material is not routinely available for immunocytochemistry. In spite of these limitations, there were few major discrepancies. These might be reduced if pathologist aspirators were able to perform FNAs and collect material for further studies, where necessary. This would allow identification of medullary carcinomas and malignant lymphomas.
Conclusion  FNA of thyroid lesions is a useful investigation in our clinical setting, however, some areas of potential for improvement have been identified.