Apoptosis is the most efficient death-pathway in tumor cells after topoisomerase II inhibition

To compare the efficiency of apoptosis and other modes of cell death in killing tumor cells after the induction of DNA damage by topoisomerase inhibitors like etoposide. This study was carried out in the Tumor Biology Department, National Cancer Institute, Cairo University, Cairo, Egypt, from September 2005 to August 2007. The breast cancer MCF7, the cervix carcinoma, human cervical adenocarcinoma [Hela], and the brain tumor U251 cell lines were exposed to etoposide. Apoptosis was detected using the flow cytometry and the DNA ladder formation methods. Cell viability was determined by a colorimetric assay, and the residual DNA double-strand breaks [dsb] were measured by gel electrophoresis. The Hela cells were the most, the MCF7's were moderately, whereas the U251's were the least sensitive to etoposide. Apoptosis was detected only in Hela cells whereas the other 2 cell lines showed a very low level of apoptosis [only 3% increase above the control cells]. At equitoxic drug concentrations [namely IC50], the Hela cells showed the lowest amount of non-repaired DNA dsb, and the MCF7's showed the highest amount, whereas the U251 cells showed a moderate amount. These results indicate that although other modes of cell death exist, apoptosis is the most efficient and requires lower drug concentrations and fewer numbers of non-repaired dsb to give the same killing effect. Clinically, this means that tumors that can execute apoptosis may require lower doses of topoisomerase inhibitors than those that lost the ability to exercise apoptosis

clinical significance of real-time PCR for rapid diagnosis of mycoplasma pneumoniae in respiratory tract infection

Mycoplasma pneumoniae [M. pneumoniae] is a common cause of respiratory tract infections [RTIs]. Diagnosis of M. pneumoniae infection relies mainly on laboratory tests, as the clinical presentation is not significantly different from that seen with other pathogens causing RTI. Diagnosis has traditionally been obtained by serological diagnosis, but increasingly, molecular techniques have been applied. However, the number of studies actually comparing these assays is limited. The Real Time PCR [RT-PCR] assay for detection of M. pneumoniae was used and comparing with a conventional PCR assay, and with serology using IgM Immunofluorescence assay [IFA]. The study included: 70 adult patients with manifestations of RTIs and 20 age matched healthy controls. All patients and controls subjected to the following: thorough history and clinical examination, routine investigations as complete blood count [CBC], erythrocyte sedimentation rate [ESR], C-reactive protein [CRP], and chest-X-ray, blood samples were collected for serological testing [IgM-IFA]. nasopharyngeal swabs were done for PCR assays and for culture techniques. Diagnosis of M. pneumoniae was made on the basis of tbe conventional PCR results. The conventional PCR was positive for M. pneumoniae in 18 patients [25.7%] out of overall 70 patients and all the 18 patients positive by conventional PCR were also positive by RT-PCR. While, all controls were PCR negative by both techniques. The nasopharyngeal swab culture for M. pneumoniae was positive in 10[14.3%] out of 70 patients and negative in all 20 healthy controls. However, the M. pneumoniae IgM-IFA was positive for 21 [30%] out of 70 patients. So IgM IFA was positive in 3 patients with negative PCR, However, one case was positive for IgM- IFA among healthy controls. Comparing to conventional PCR, the RT-PCR gives 100% sensitivity, 100% specificity and 100% accuracy for diagnosis of M. pneumoniae, followed by IgM- IFA that give a sensitivity of 100%, specificity of 94.2% and accuracy of 95.7%, and the culture of M. pneumoniae gives 55.6% for sensitivity, 100% for specificity and 88.5% for diagnostic accuracy. The comparison of clinical data of patients diagnosed with M. pneumoniae infection [+ve PCR] and those who were M. pneumoniae negative [-ye PCR] revealed that no significant difference was reported as regard age or disease duration between the two groups of patients, while rhinitis was significantly more prevalent in the Mycoplasma-negative patients [p<0.01]. No significant difference was seen between the two groups regarding headache, malaise, sore throat, vomiting, fever, wheeze, or chest radiographic infiltrate [p>0.05]. Also, WBC counts, CRP levels and ESR values were significantly increased in PCR positive group as comparing to PCR negative group [p<0.05]. The molecular methods are superior for diagnosis of M. pneumoniae regarding accuracy, and providing more rapid diagnosis. In addition, using Real-Time PCR assay involves less hands, short time for diagnosis and meanwhile, rapid treatment and monitoring therapy