Circulating tumor cells (CTCs) play a significant role in cancer diagnosis and personalized therapy, and it is still a significant challenge to efficiently capture and gently release CTCs from clinical samples for downstream manipulation and molecular analysis. Many CTC devices incorporating various nanostructures have been developed for CTC isolation with sufficient capture efficiency, however, fabricating such nanostructured substrates often requires elaborate design and complicated procedures. Here we fabricate a degradable zinc-phosphate-based hierarchical nanosubstrate (HZnPNS), and we demonstrate its excellent CTC-capture performance along with effective cell-release capability for downstream molecular analysis. This transparent hierarchical architecture prepared by a low-temperature hydrothermal method, enables substantially enhanced capture efficiency and convenient imaging. Biocompatible sodium citrate could rapidly dissolve the architecture at room temperature, allowing that 88 ± 4% of captured cells are gently released with a high viability of 92 ± 1%. Furthermore, antiepithelial cell adhesion molecule antibody functionalized HZnPNS (anti-EpCAM/HZnPNS) was successfully applied to isolate CTCs from whole blood samples of cancer patients, as well as release CTCs for global DNA methylation analysis, indicating it will serve as a simple and reliable alternative platform for CTC detection.

Despite implications of persistence of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) in the development of hepatocellular carcinoma (HCC), little is known about serum cccDNA in HBV-infected diseases. We developed a cccDNA-selective droplet digital PCR (ddPCR) to assess cccDNA content and dynamics across different stages of HCC development. One hundred forty-seven serum samples and 35 formalin-fixed, paraffin-embedded tumor tissues were derived from patients with HCC or HBV hepatitis/cirrhosis. After specific amplification and selective digestion, probe-based ddPCR was used to quantify cccDNA copy numbers in single cells and clinical samples. The cccDNA in single HepG2.2.15 cells ranged from 0 to 10.8 copies/cell. Compared with non-HCC patients, HCC patients showed a higher cccDNA-positive rate (89.9% versus 53.2%; P = 4.22 × 10) and increased serum cccDNA contents (P = 0.002 and P = 0.041 for hepatitis and cirrhosis patients, respectively). Serum cccDNA ranged from 84 to 1.07 × 10 copies/mL. Quantification of serum cccDNA and HBV-DNA was an effective way to discriminate HCC patients from non-HCC patients, with areas under the curve of receiver operating characteristic of 0.847 (95% CI, 0.759-0.935; sensitivity, 74.5%; specificity, 93.7%). cccDNA-selective ddPCR is sensitive to detect cccDNA in single cells and different clinical samples. Combined analysis of serum cccDNA and HBV-DNA may be a promising strategy for HBV-induced HCC surveillance and antiviral therapy evaluation.

Background Accumulating evidence suggests that epigenetic changes play key roles in the pathogenesis of type 2 diabetes mellitus (T2DM). However, the dynamic regulation of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) in diabetic peripheral blood DNA remains to be elucidated. Results We collected fasting blood samples (104 patients and 108 healthy controls) and glucose-stimulated blood samples at different time points (11 patients and 5 healthy controls underwent oral glucose tolerance test (OGTT)), as well as blood samples from six couples of diabetic and control rats. A HPLC-MS/MS system was used for quantifying global 5mC and 5hmC in genomic DNA from white blood cells (WBCs), and qPCR was performed for detecting mRNA expression of SIRT6 and TETs . We found that global 5mC decreased, while global 5hmC increased in both patients and diabetic rats, with lower 5mC being a risk factor of T2DM (OR = 0.524, 95%CI 0.402–0.683, p = 1.64 × 10 −6 ). The OGTT data from patients showed that 5mC declined within 1 h and then returned to the fasting status at 2 h, while 5hmC rose from 0.5 h to 3 h with increasing glucose. However, the similar patterns were not found in the controls. The mRNA expression of TET2 , TET3 , and SIRT6 was upregulated in patients ( p = 0.012, p = 0.026, and p = 0.035, respectively). The similar results were observed in diabetic OGTT and rats. Correlation analysis indicated that SIRT6 was positively correlated with TET2 in humans ( r = 0.277, p < 0.001) and rats ( r = 0.942, p < 0.001), in addition to a correlation between glucose and SIRT6 ( r = 0.162, p = 0.045) and TET2 ( r = 0.174, p = 0.036). Conclusions Hyperglycemia appeared to promote the mRNA expression of SIRT6 and TETs , which in turn might cause the dynamic changes of 5mC and 5hmC in WBCs from T2DM patients. Electronic supplementary material The online version of this article (10.1186/s13148-019-0660-y) contains supplementary material, which is available to authorized users.

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