Salvage neck surgery in recurrent nodal NPC: Do all patients require a comprehensive neck dissection in the modern MRI era?

OBJECTIVE: The standard treatment for regional failure in nasopharyngeal carcinoma (NPC) is the radical neck dissection (RND). Our study sought to determine if magnetic resonance imaging (MRI) may accurately predict nodal involvement to allow selected levels of neck dissection to be preserved. STUDY DESIGN AND SETTING: We analysed retrospectively all NPC patients in our centre undergoing neck dissections as salvage therapy for nodal recurrence. Nodal involvement based on the preoperative MRI was assessed and compared with postoperative histopathology. METHODS: This is a retrospective study conducted on patients in our centre with recurrent NPC from February 2002 to February 2017. Patients were identified from the database of the otolaryngology oncology division at our institution. Of these, 28 patients met all our inclusion and exclusion criteria. We calculated sensitivity and specificity as well as average number of nodes per patient. RESULTS: In our study, we calculated the false negative and false positive rates of preoperative MRI neck by levels. Overall sensitivity of MRI picking up disease by level was 76% and specificity was 86%. CONCLUSION: Based on our study, we will be missing a total of 10 (7.1%) diseased neck levels in eight (28.5%) patients. MRI alone, therefore, does not provide enough information to allow safe selective preservation of neck levels in surgical salvage of neck recurrences in NPC.

[A pre-conception cohort study of fertility and its related factors among couples with the intention of conception].

Objective: To describe fertility and explore factors associated with it among pre-conception couples of childbearing age. Methods: Based on the pre-conceptional offspring trajectory study of the School of Public Health of Fudan University, couples of childbearing age who participated in the pre-conception physical examination in Shanghai Jiading District from 2016 to 2021 were recruited and followed up. Couples' time to pregnancy (TTP) was analyzed and Cox proportional hazards regression model was used to explore the factors associated with TTP. Kaplan-Meier was used to calculate each menstrual cycle's cumulative pregnancy rate. Results: A total of 1 095 preconception couples were included in the analysis, the M(Q1,Q3)of TTP was 4.33 (2.41, 9.78) menstrual cycles. Age of women (FR=0.90, 95%CI: 0.85-0.95, P<0.001), women who were overweight or obese before pregnancy (FR=0.36, 95%CI: 0.24-0.55, P<0.001), women who were exposed to second-hand smoking (FR=0.63, 95%CI: 0.44-0.92, P=0.016), women whose home or office had been renovated in the past 2 years and had a particular smell (FR=0.46, 95%CI: 0.26-0.81, P=0.008) were risk factors for impaired fertility. Regular menstrual cycles (FR=1.64, 95%CI: 1.16-2.31, P=0.005), females who often drank tea/coffee (FR=1.55, 95%CI: 1.11-2.17, P=0.011) and males who took folic acid before conception (FR=2.35, 95%CI: 1.38-4.23, P=0.002) were associated with better fertility. The cumulative pregnancy rate of 3, 6, and 12 menstrual cycles was 37.6%, 64.4%, and 78.4%, respectively. Conclusion: Older couples, overweight or obesity before pregnancy, irregular menstruation, exposure to secondhand smoke and decoration pollutants in females are associated with impaired fertility. Frequent tea/coffee drinking before pregnancy in females and taking folic acid before pregnancy in males are associated with shortened conception time.

Genetic diversity and structure of the threatened species Sinopodophyllum hexandrum (Royle) Ying.

Sinopodophyllum hexandrum is an important medicinal plant that has been listed as an endangered species, making the conservation of its genetic diversity a priority. Therefore, the genetic diversity and population structure of S. hexandrum was investigated through inter-simple sequence repeat analysis of eight natural populations. Eleven selected primers generated 141 discernible fragments. The percentage of polymorphic bands was 37.59% at the species level, and 7.66-24.32% at the population level. Genetic diversity of S. hexandrum was low within populations (average HE = 0.0366), but higher at the species level (HE = 0.0963). Clear structure and high genetic differentiation were detected between populations using unweighted pair groups mean arithmetic and principle coordinate analysis. Clustering approaches clustered the eight sampled populations into three major groups, and AMOVA confirmed there to be significant variation between populations (63.27%). Genetic differentiation may have arisen through limited gene flow (Nm = 0.3317) in this species. Isolation by distance among populations was determined by comparing genetic distance versus geographical distance using the Mantel test. The results revealed no correlation between spatial pattern and geographic location. Given the low within-population genetic diversity, high differentiation among populations, and the increasing anthropogenic pressure on this species, in situ conservation measures, in addition to sampling and ex situ preservation, are recommended to preserve S. hexandrum populations and to retain their genetic diversity.

BCL-2 expression delays drug-induced apoptosis but does not increase clonogenic survival after drug treatment in HeLa cells.

Apoptosis is a major form of cell death induced by chemotherapeutic drugs. Overexpression of the proto-oncogene bcl-2 can prevent apoptosis in various types of cells. We have constructed a HeLa S3 cell line in which the expression of bcl-2 can be controlled by the concentration of tetracycline in the medium. Using this system, we show that apoptosis induced by various cytostatic treatments could be delayed by the overexpression of bcl-2, as assayed by vital dye exclusion, apoptotic nuclei morphology, DNA histogram shift, and DNA fragmentation. Quantitative analysis revealed a hyperbolic curve when protection from apoptosis was plotted against the amount of Bcl-2. When cells were treated with aphidicolin for 12, 24, or 36 h and then replated in fresh media to assay for colony formation, the majority of cells that did not show apoptotic morphology at the time of drug removal failed to form colonies. Furthermore, Bcl-2 did not increase colony formation after 12-36 h of aphidicolin treatment. Therefore, with aphidicolin treatment, cells were committed to the death program upstream of the point of Bcl-2 action.

Inhibition of apoptosis by overexpressing Bcl-2 enhances gene amplification by a mechanism independent of aphidicolin pretreatment.

To study the effect of apoptosis on gene amplification, we have constructed HeLa S3 cell lines in which the expression of bcl-2 (BCL2) can be controlled by tetracycline in the growth medium. Induction of Bcl-2 expression caused a temporary delay of apoptosis and resulted in roughly a 3-fold increase in the frequency of resistant colonies when cells were selected with trimetrexate. This resistance was due to amplification of the dihydrofolate reductase gene. Cells grown out of the pooled resistant colonies retained the same level of resistance to trimetrexate whether Bcl-2 was induced or repressed, consistent with the theory that Bcl-2 functions by facilitating gene amplification, rather than being the resistance mechanism per se. Pretreating cells with aphidicolin is another method to increase gene amplification frequency. When Bcl-2-expressing cells were pretreated with aphidicolin, the resulting increase in gene amplification frequency was approximately the product of the increases caused by aphidicolin pretreatment or Bcl-2 expression alone, indicating that Bcl-2 increases gene amplification through a mechanism independent of that of aphidicolin pretreatment. These results are consistent with the concept that gene amplification occurs at a higher frequency during drug-induced cell cycle perturbation. Bcl-2 evidently increases the number of selected amplified colonies by prolonging cell survival during the perturbation.

Tetracycline-controlled gene expression system achieves high-level and quantitative control of gene expression.

The tetracycline-controlled gene expression system utilizes the control elements of the tetracycline resistance operon encoded in TnlO of Escherichia coli to control gene expression in eukaryotic cells. Here we demonstrate the quantitative control of the expression of the luciferase gene, dihydrofolate reductase gene, and bcl-2 gene in HeLa S3 or Chinese hamster ovary AA8 cells using the tetracycline-controlled gene expression system. Regardless of the host cell lines or the genes being expressed, there is a common range of tetracycline concentration within which the expression of genes is most sensitively regulated. In addition, the maximal gene expression level of the tetracycline-controlled gene expression system is higher than that of the wild-type CMV promoter/enhancer-driven system. Nonetheless, careful selection of stably transfected clones is necessary to achieve the optimally regulated gene expression using this system.