Decrease of spermatozoa with an unbalanced chromosome content after cell sorting in men carrying a structural chromosomal abnormality.

BACKGROUND: We showed that in men with a constitutional chromosomal abnormality, DNA fragmentation was significantly higher in chromosomally unbalanced spermatozoa than in spermatozoa with a normal or balanced chromosomal content. These results could be explained by a phenomenon already described in infertile men: abortive apoptosis. OBJECTIVES: To determine whether magnetic-activated cell separation could select spermatozoa with lower levels of DNA fragmentation and unbalanced chromosome content in men carrying a structural chromosomal abnormality. MATERIALS AND METHODS: The spermatozoa of ten males with a chromosomal rearrangement were separated into two populations using magnetic-activated cell separation (annexin V (-) and annexin V (+) fractions), in order to study meiotic segregation by fluorescence in situ hybridization, the percentage of spermatozoa with an externalization of phosphatidylserine by annexin V staining and DNA fragmentation by TdT-mediated dUTP nick-end labeling on the whole ejaculate and on selected spermatozoa in the same patient. RESULTS: For all patients, the percentage of spermatozoa with externalization of phosphatidylserine decreased in the annexin V (-) fraction and increased in the annexin V (+) fraction as compared to the frozen-thawed semen sample. The rates of DNA fragmentation were statistically much lower in the annexin V (-) fraction when compared to the rate before magnetic-activated cell separation for all but one patient. Conversely, we observed a statistically significantly higher rate of DNA fragmentation in the annexin V (+) fraction for six patients. After magnetic-activated cell separation, there was a significant increase of normal/balanced spermatozoa in the fraction of annexin V (-) for all patients. Conversely, we observed a significant decrease in the fraction of annexin V (+) for seven patients. DISCUSSION AND CONCLUSIONS: Magnetic-activated cell separation is a promising tool for increasing the selection of healthy spermatozoa, with a decrease in the number of spermatozoa with externalization of phosphatidylserine, DNA fragmentation, and chromosome unbalance, for use in assisted reproductive technologies such as intracytoplasmic sperm injection for males with a chromosomal structural abnormality.

Gemcitabine plus sirolimus for relapsed and progressing osteosarcoma patients after standard chemotherapy: a multicenter, single-arm phase II trial of Spanish Group for Research on Sarcoma (GEIS).

BACKGROUND: Patients with relapsed unresectable osteosarcoma represents an unmet need, so active and safe systemic treatments are required. Fas cell surface death receptor and mammalian target of rapamycin pathways are implicated in progressing osteosarcoma, and we had preclinical and clinical experience with a scheme that targets both pathways. Therefore, we designed a phase II trial with gemcitabine plus rapamycin, to determine the efficacy and safety, in this subset of patients. PATIENTS AND METHODS: A multicenter, single-arm phase II trial was sponsored by the Spanish Group for Research on Sarcoma. Osteosarcoma patients, relapsed or progressing after standard chemotherapy and unsuitable for metastasectomy received gemcitabine and rapamycin p.o. 5 mg/day except for the same day of gemcitabine administration, and the day before. The main end point was 4-month progression-free survival rate (PFSR), with the assumption that rates higher than 40% would be considered as an active regimen. Translational research aimed to correlate biomarkers with the clinical outcome. RESULTS: Thirty-five patients were enrolled and received at least one cycle. PFSR at 4 months was 44%, and after central radiologic assessment, 2 partial responses and 14 stabilizations (48.5%) were reported from 33 assessable patients. The most frequent grade 3-4 adverse events were: neutropenia (37%), thrombocytopenia (20%), anemia (23%), and fatigue (15%); however, only three patients had febrile neutropenia. Positive protein expression of RRM1 significantly correlated with worse PFS and overall survival, while positivity of P-ERK1/2 was correlated with significant better overall survival. CONCLUSION: Gemcitabine plus sirolimus exhibits satisfactory antitumor activity and safety in this osteosarcoma population, exceeding the prespecified 40% of 4-month PFSR. The significant correlation of biomarkers with clinical outcome encourages further prospective investigation.

Health-related quality of life is associated with COPD severity: a comparison between the GOLD staging and the BODE index.

Chronic obstructive pulmonary disease (COPD) is a highly prevalent disease. Currently, severity Global initiative for chronic Obstructive Lung Disease (GOLD) criteria are used to diagnose the severity of COPD, but a new grading system, the body mass index, bronchial obstruction, dyspnea, exercise (BODE) index, was recently proposed to provide useful prognostic information. The objective of this study is to evaluate the association between health-related quality of life (HRQOL) and COPD severity assessed by two criteria: the GOLD classification and the BODE index. Sixty-four patients with COPD were examined with lung function tests and specific and generic HRQOL questionnaires (St. George's Respiratory Questionnaire [SGRQ], Nottingham Health Profile [NHP]). Participants were divided into four severity groups using the GOLD guidelines and the BODE index quartiles. The association between NHP and SGRQ subscales, and the BODE index was significant (P < 0.01). However, the GOLD classification shows a correlation only with SGRQ total score (P < 0.05) but not with NHP or SGRQ subscales. There was an association of the SGRQ total score between the severity groups of BODE (P = 0.0001), but there was no difference in the SGRQ total score between the severity groups of GOLD classification (P = 0.244). The present study suggests that COPD severity assessed by the BODE index can be more directly related with HRQOL.

Quality of life in patients with chronic obstructive pulmonary disease: the predictive validity of the BODE index.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is currently the fourth cause of mortality and morbility in the developed world. Patients with COPD experience a progressive deterioration of health-related quality of life (HRQOL). A new model of severity classification, the body mass index, bronchial obstruction, dyspnoea, exercise (BODE) index, has recently been proposed. OBJECTIVE: To evaluate the relationship between HRQOL and the BODE index, and the predictive ability of BODE on HRQOL measurements. METHODS: Two HRQOL questionnaires were administered, namely the Nottingham Health Profile (NHP) and St George's Respiratory Questionnaire (SGRQ), in a sample of 67 patients with severe COPD. RESULTS: Pearsons correlation coefficient analysis shows a positive correlation between the BODE index and the total scores of the specific (P < 0.001), and general HRQOL (P < 0.001); the analysis shows a significant correlation between the BODE index and the subscales of symptoms, activity and impact of SGRQ (P < 0.001) and the subscales energy and physical mobility of the NHP (P < 0.001). The regression analysis shows that the BODE index is a significant predictor of HRQOL, explaining 46,1% of the total score of the SGRQ (P < 0.001) and 14.8% of the total score of the NHP (P < 0.001). CONCLUSIONS: The BODE index is good at predicting the worsening of HRQOL in patients with severe COPD.

Conditional expression of RNase P in the cyanobacterium Synechocystis sp. PCC6803 allows detection of precursor RNAs. Insight in the in vivo maturation pathway of transfer and other stable RNAs.

We have constructed a strain (CT1) that expresses RNase P conditionally with the aim to analyze the in vivo tRNA processing pathway and the biological role that RNase P plays in Synechocystis 6803. In this strain, the rnpB gene, coding for the RNA subunit of RNase P, has been placed under the control of the petJ gene promoter (P(petJ)), which is repressed by copper, cell growth, and accumulation of RNase P RNA is inhibited in CT1 after the addition of copper, indicating that the regulation by copper is maintained in the chimerical P(petJ)-rnpB gene and that RNase P is essential for growth in Synechocystis. We have analyzed several RNAs by Northern blot and primer extension in CT1. Upon addition of copper to the culture medium, precursors of the mature tRNAs are detected. Furthermore, our results indicate that there is a preferred order in the action of RNase P when it processes a dimeric tRNA precursor. The precursors detected are 3'-processed, indicating that 3' processing can occur before 5' processing by RNase P. The size of the precursors suggests that the terminal CCA sequence is already present before RNase P processing. We have also analyzed other potential RNase P substrates, such as the precursors of tmRNA and 4.5 S RNA. In both cases, accumulation of larger than mature size RNAs is observed after transferring the cells to a copper-containing medium.