Applying massively parallel sequencing to paternity testing on the Ion Torrent Personal Genome Machine

Massively parallel sequencing (MPS) is a promising supplementary method for forensic genetics and has gradually been applied to forensic casework. In this study, we applied MPS to forensic casework on an Ion Torrent Personal Genome Machine to evaluate its performance in paternity testing with mismatched STR loci. A total of 15 samples from seven cases containing one mismatched locus by capillary electrophoresis typing were analyzed. Combined paternity index (CPI) and relative chance of paternity were calculated according to the International Society for Forensic Genetics guidelines and the Chinese national standards recommended for paternity testing. With simultaneous analysis of enough STR loci, the results support the certainty of paternity, and the mismatched alleles were considered to be mutations (CPI > 10,000). With the detection of allele sequence structures, the origins of the mutations were inferred in some cases. Meanwhile, nine STRs (CSF1PO, D1S1656, D2S441, D2S1338, D3S1358, D8S1179, D12S391, D21S11 and D4S2408) were found in an increased number of unique alleles and three new alleles in three STRs (D2S441, D21S11, and FGA) that have not been reported before were detected. Therefore, MPS can provide valuable information for forensic genetics research and play a promising role in paternity testing.     

Evaluation of the in situ assay for HBV DNA: An observational real-world study in chronic hepatitis B

The visualization of intrahepatic hepatitis B virus (HBV) DNA by in situ hybridization (ISH) has uncovered some interesting aspects of HBV life cycle at the single-cell level. In the current study, we intend to evaluate the reliability and robustness of this assay in the real-world clinical scenario and its relationship with currently available clinical biomarkers in chronic hepatitis B (CHB) patients.In this cross-sectional study, 94 CHB patients and 10 patients with non-HBV related liver diseases were enrolled. Liver biopsies and routine histopathology analysis were performed. Intrahepatic HBV DNA and viral antigens (HBsAg and HBcAg) were detected by ISH and immunohistochemistry (IHC), respectively. The basic biochemical and virological parameters such as alanine transaminase, serum HBV DNA, and serum HBsAg were measured.The HBV DNA-ISH assay showed 55.8% (53/94 cases) positive rate in CHB patients, no false positive was found in non-HBV related hepatitis. The IHC of HBsAg and HBcAg showed a positive rate of 94.7% (89/94 cases) and 19.5% (17/87 cases), respectively. Quantification of HBV DNA-ISH signal showed a significant correlation with serum HBV DNA (r_s = 0.6223, P < .0001). In addition, the staining pattern of HBV DNA in situ in the context of collagen deposition informed the histopathological progression of chronic liver disease.The application of this ISH assay in evaluating intrahepatic viral replication in real-world CHB patients showed favorable performance. It can be a complementation to conventional liver histopathology examination and IHC detection of viral antigens. This methodology provides an intuitive assessment of virological and pathological state of CHB patients, and further supports clinical diagnosis and management.     

The randomized placebo-phase design for clinical trials

Randomized controlled trials (RCTs) are the criterion standard method for evaluating the effectiveness of medical treatments. There are situations, however, where the possibility of being in the control group in a RCT is unacceptable to potential subjects or their physicians. This lack of acceptance is a reason for poor accrual. We developed and validated a new clinical trial design for survival data that may allay concerns about not receiving an investigational product and should be more acceptable. Called the randomized placebo-phase design (RPPD), this new design asks whether, on average, those subjects who begin active treatment sooner respond sooner than those who begin it later. Using Monte Carlo computer simulations, we demonstrated that the design is valid and may offer advantages over traditional RCTs in some situations. The RPPD may be especially useful when highly potent therapies for rare diseases are tested or when accrual may be otherwise difficult.     

Socioeconomic status does not affect the outcome of liver transplantation

The outcome of liver transplantation is dependent on many factors. It was suggested that racial disparities in outcome may be related to differences in socioeconomic status (SES). In this retrospective study, we analyzed the effect of SES on graft and patient survival. Two hundred seventy-six adult patients who underwent liver transplantation at our institution from July 1988 to June 2001 were included in the analysis. Educational and occupation statuses were coded using established criteria (Hollingshead Index of Social Status [HI]). SES then was calculated using the HI formula: SES = education level × 3 + occupation × 5, and categorized into four groups: group 1, score less than 29 (n = 71); group 2, score of 29 to 42 (n = 82); group 3, score of 42 to 53 (n = 69); and group 4, score greater than 53 (n = 54). Kaplan-Meier analysis was used for graft and patient survival, and Cox regression analysis was used to determine the effect of confounding factors. Demographics of all four groups were similar. One-, 2-, and 5-year graft and patient survival did not differ significantly across groups by Kaplan-Meier and Cox regression survival analysis. In conclusion, SES did not predict graft and patient survival after liver transplantation. (Liver Transpl 2002;8:1133-1137.)     

Pluripotent factor lin-28 and its homologue lin-28b in epithelial ovarian cancer and their associations with disease outcomes and expression of let-7a and IGF-II

Lin-28 and lin-28B are RNA-binding proteins which can block microRNA let-7 maturation and affect the differentiation and proliferation of embryonic stem cells. Lin-28 may also regulate the expression of insulin-like growth factor II (IGF-II). As one of the pluripotent factors involved in making induced pluripotent stem cells (iPS), lin-28 is considered a potential therapeutic target for cancer treatment. To further understand the role of lin-28 in cancer, we analysed the expression of lin-28 and its homologue lin-28B in tumour samples, and evaluated their associations with let-7a maturation, IGF-II expression, disease features and outcomes in 211 patients with primary epithelial ovarian cancer. The analysis showed that both lin-28 and lin-28B were positively correlated with primary and pre-let-7a-3; lin-28B, not lin-28, was inversely correlated with mature let-7a. A positive correlation was also observed between lin-28B and IGF-II expression, while no association was found between lin-28B and IGF-I or IGFBP-3. The study further demonstrated that lin-28B expression was associated with the risk of disease progression and death; patients with high lin-28B had shorter progression-free and overall survival than those with low lin-28B. These results seem to support the findings of recent in vitro experiments, showing that lin-28 blocks the process of let-7a maturation. Our study also suggests that lin-28B may promote ovarian cancer progression and serve as an unfavourable prognostic marker for the disease. The correlation between lin-28B and IGF-II indicates that the growth factor may mediate the effect of lin-28B on tumour growth.