Antimicrobial peptaibols, novel suppressors of tumor cells, targeted calcium-mediated apoptosis and autophagy in human hepatocellular carcinoma cells

<p>Abstract</p> <p>Background</p> <p>Hepatocellular carcinoma (HCC) is one of the most common cancers in the world which is highly chemoresistant to currently available chemotherapeutic agents. Thus, novel therapeutic targets are needed to be sought for the successful treatment of HCC. Peptaibols, a family of peptides synthesized non-ribosomally by the <it>Trichoderma </it>species and other fungi, exhibit antibiotic activities against bacteria and fungi. Few studies recently showed that peptaibols exerted cytotoxicity toward human lung epithelial and breast carcinoma cells. However, the mechanism involved in peptaibol-induced cell death remains poorly understood.</p> <p>Results</p> <p>Here, we showed that Trichokonin VI (TK VI), a peptaibol from <it>Trichoderma pseudokoningii </it>SMF2, induced growth inhibition of HCC cells in a dose-dependent manner. It did not obviously impair the viability of normal liver cells at lower concentration. Moreover, the suppression of cell viability resulted from the programmed cell death (PCD) with characteristics of apoptosis and autophagy. An influx of Ca²⁺triggered the activation of μ-calpain and proceeded to the translocation of Bax to mitochondria and subsequent promotion of apoptosis. On the other hand, typically morphological characteristics consistent with autophagy were also observed by punctate distribution of MDC staining and the induction of LC3-II, including extensive autophagic vacuolization and enclosure of cell organelles by these autophagosomes. More significantly, specific depletion of Bak expression by small RNA interfering (siRNA) could partly attenuate TK VI-induced autophagy. However, siRNA against Bax led to increased autophagy.</p> <p>Conclusion</p> <p>Taken together, these findings showed for the first time that peptaibols were novel regulators involved in both apoptosis and autophagy, suggesting that the class of peptaibols might serve as potential suppressors of tumor cells.</p

High persistence rate of hepatitis B virus in a hydrodynamic injection-based transfection model in C3H/HeN mice

AIM: To optimize the viral persistence rate in a hydrodynamic injection (HI) based hepatitis B virus (HBV) transfection mouse model.METHODS: (1) 5-6-wk-old male C3H/HeN and C57BL/6 mice were hydrodynamically injected with 10 μg endotoxin-free pAAV/HBV1.2 plasmid DNA via the tail vein. Hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg) and HBV DNA, both in the serum and liver, were detected at different time points post HI by ELISA, immunohistochemical staining or quantitative polymerase chain reaction (PCR); (2) male C3H/HeN and C57BL/6 mice, either hydrodynamically injected mice at 10 wk post HI or naïve mice, were all immunized subcutaneously with 5 μg HBsAg formulated in complete Freund's adjuvant three times at a 2-wk interval. Two weeks after the final immunization, splenocytes were isolated for T cell function analysis by ELISPOT assay; and (3) five weeks post HI, C3H/HeN mice were intragastrically administered 0.1 mg/kg entecavir once a day for 14 d, or were intraperitoneally injected with 1 mg/kg interferon (IFN)-α twice a week for 2 wk, or were treated with PBS as controls. The sera were collected and assayed for HBV DNA on days 0, 7 and 14 after drug treatment.RESULTS: (1) Approximately 90% (22/25) of the injected C3H/HeN mice were still HBsAg-positive at 46 wk post HI, whereas HBsAg in C57BL/6 mice were completely cleared at 24 wk. Serum levels of HBeAg in C3H/HeN mice were higher than those in C57BL/6 mice from 4 wk to 46 wk. HBV DNA levels in the hydrodynamically injected C3H/HeN mice were higher than those in the C57BL/6 mice, both in the serum (from 4 wk to 46 wk) and in the liver (detected at 8 wk and 46 wk post HI). Histology showed that hepatitis B core antigen and HBsAg were expressed longer in the liver of C3H/HeN mice than in C57BL/6; (2) HBsAg specific T cell responses after HBsAg vaccination in hydrodynamically injected C3H/HeN and C57BL/6 mice, or naive control mice were detected by ELISPOT assay. After stimulation with HBsAg, the frequencies of IFN-γ producing splenocytes in the hydrodynamically injected C3H/HeN mice were significantly lower than those in hydrodynamically injected C57BL/6 mice, control C3H/HeN and control C57BL/6 mice, which were 0, 17 ± 7, 18 ± 10, and 41 ± 10 SFCs/10(6) splenocytes, respectively, and the mean spot sizes showed the same pattern. Even just stimulated with PMA and ionomysin, T-cell responses elicited in the vaccinated control C3H/HeN were much higher than those in hydrodynamically injected C3H/HeN mice; and (3) For drug treatment experiments on the hydrodynamically injected C3H/HeN mice, serum HBV DNA levels in the entecavir treatment group declined (131.2 folds, P &lt; 0.01) on day 7 after treatment and kept going down. In the group of IFN-α treatment, serum HBV DNA levels declined to a lowest point (6.42 folds, P &lt; 0.05) on 7 d after treatment and then rebounded.CONCLUSION: We have developed a novel HI-based HBV transfection model using C3H/HeN mice, which had a higher HBV persistence rate than the classic C57BL/6 mouse model.</p

Safety and efficacy of Hypofractionated stereotactic radiosurgery for high-grade Gliomas at first recurrence: a single-center experience.

BACKGROUND: The optimal treatment for recurrent high-grade gliomas (rHGGs) remains uncertain. This study aimed to investigate the efficacy and safety of hypofractionated stereotactic radiosurgery (HSRS) as a first-line salvage treatment for in-field recurrence of high-grade gliomas. METHODS: Between January 2016 and October 2019, 70 patients with rHGG who underwent HSRS were retrospectively analysed. The primary endpoint was overall survival (OS), and secondary endpoints included both progression-free survival (PFS) and adverse events, which were assessed according to Common Toxicity Criteria Adverse Events (CTCAE) version 5. The prognostic value of key clinical features (age, performance status, planning target volume, dose, use of bevacizumab) was evaluated. RESULTS: A total of 70 patients were included in the study. Forty patients were male and 30 were female. Forty-nine had an initial diagnosis of glioblastoma (GBM), and the rest (21) were confirmed to be WHO grade 3 gliomas. The median planning target volume (PTV) was 16.68 cm3 (0.81–121.96 cm3 ). The median prescribed dose was 24 Gy (12–30 Gy) in 4 fractions (2–6 fractions). The median baseline of Karnofsky Performance Status (KPS) was 70 (40–90). With a median follow-up of 12.1 months, the median overall survival after salvage treatment was 17.6 months (19.5 and 14.6 months for grade 3 and 4 gliomas, respectively; p = .039). No grade 3 or higher toxicities was recorded. Multivariate analysis showed that concurrent bevacizumab with radiosurgery and KPS \u3e 70 were favourable prognostic factors for grade 4 patients with HGG. CONCLUSIONS: Salvage HSRS showed a favourable outcome and acceptable toxicity for rHGG. A prospective phase II study (NCT04197492) is ongoing to further investigate the value of hypofractionated stereotactic radiosurgery (HSRS) in rHGG

Charge-changing cross section measurements of 300 MeV/nucleon 28^{28}Si on carbon and data analysis

Charge-changing cross section ($\sigma_{\text{cc}}$) measurements via the transmission method have made important progress recently aiming to determine the charge radii of exotic nuclei. In this work, we report a new $\sigma_{\text{cc}}$ measurement of 304(9) MeV/nucleon $^{28}$Si on carbon at the second Radioactive Ion Beam Line in Lanzhou (RIBLL2) and describe the data analysis procedure in detail. This procedure is essential to evaluate the systematic uncertainty in the transmission method. The determined $\sigma_{\mathrm{cc}}$ of 1125(11) mb is found to be consistent with the existing data at similar energies. The present work will serve as a reference in the $\sigma_{\text{cc}}$ determinations at RIBLL2.Comment: 9 pages, 13 figures, to be published in Chinese Physics

Upregulation of Circulating PD-L1/PD-1 Is Associated with Poor Post-Cryoablation Prognosis in Patients with HBV-Related Hepatocellular Carcinoma

BACKGROUND: The programmed cell death-1 receptor/programmed cell death-1 ligand (PD-1/PD-L1) pathway plays a crucial role in tumor evasion from host immunity. This study was designed to evaluate the association between circulating PD-L1/PD-1 and prognosis after cryoablation in patients with HBV-related hepatocellular carcinoma (HCC). METHODOLOGY/PRINCIPAL FINDINGS: In the present study, 141 HBV-related HCC patients were enrolled and of those 109 patients received cryoablation. Circulating PD-L1/PD-1 expression was tested by flow cytometry, and 23 patients were simultaneously evaluated for intratumoral PD-L1 expression by immunohistochemical staining. Circulating PD-1/PD-L1 expression was associated with severity of diseases in patients with HCC, and the circulating PD-L1 expression was closely correlated with intratumoral PD-L1 expression. Of the clinical parameters, PD-1/PD-L1 expression was associated with tumor size, blood vessel invasion and BCLC staging. Moreover, PD-1/PD-L1 expression dropped after cryoablation while being elevated at the time of tumor recurrence. Patients with higher expression of circulating PD-L1, as well as circulating PD-1, had a significantly shorter overall survival and tumor-free survival than those with lower expression. Multivariate analysis confirmed that circulating PD-L1 could serve as an independent predictor of overall survival and tumor-recurrence survival in HCC patients after cryoablation. CONCLUSIONS/SIGNIFICANCE: Upregulation of circulating PD-L1/PD-1 is associated with poor post-cryoablation prognosis in patients with HBV-related hepatocellular carcinoma

Genotype V Japanese Encephalitis Virus Is Emerging

Japanese encephalitis (JE) is a global public health issue that has spread widely to more than 20 countries in Asia and has extended its geographic range to the south Pacific region including Australia. JE has become the most important cause of viral encephalitis in the world. Japanese encephalitis viruses (JEV) are divided into five genotypes, based on the nucleotide sequence of the envelope (E) gene. The Muar strain, isolated from patient in Malaya in 1952, is the sole example of genotype V JEV. Here, the XZ0934 strain of JEV was isolated from Culex tritaeniorhynchus, collected in China. The complete nucleotide and amino acid sequence of XZ0934 strain have been determined. The nucleotide divergence ranged from 20.3% to 21.4% and amino acid divergence ranged from 8.4% to 10.0% when compared with the 62 known JEV isolates that belong to genotype I–IV. It reveals low similarity between XZ0934 and genotype I–IV JEVs. Phylogenetic analysis using both complete genome and structural gene nucleotide sequences demonstrates that XZ0934 belongs to genotype V. This, in turn, suggests that genotype V JEV is emerging in JEV endemic areas. Thus, increased surveillance and diagnosis of viral encephalitis caused by genotype V JEV is an issue of great concern to nations in which JEV is endemic

Genomic and oncogenic preference of HBV integration in hepatocellular carcinoma

Hepatitis B virus (HBV) can integrate into the human genome, contributing to genomic instability and hepatocarcinogenesis. Here by conducting high-throughput viral integration detection and RNA sequencing, we identify 4,225 HBV integration events in tumour and adjacent non-tumour samples from 426 patients with HCC. We show that HBV is prone to integrate into rare fragile sites and functional genomic regions including CpG islands. We observe a distinct pattern in the preferential sites of HBV integration between tumour and non-tumour tissues. HBV insertional sites are significantly enriched in the proximity of telomeres in tumours. Recurrent HBV target genes are identified with few that overlap. The overall HBV integration frequency is much higher in tumour genomes of males than in females, with a significant enrichment of integration into chromosome 17. Furthermore, a cirrhosis-dependent HBV integration pattern is observed, affecting distinct targeted genes. Our data suggest that HBV integration has a high potential to drive oncogenic transformation

Targeting tumor-associated macrophages by anti-tumor Chinese materia medica

Tumor-associated macrophages (TAMs) play a key role in all stages of tumorigenesis and tumor progression. TAMs secrete different kinds of cytokines, chemokines, and enzymes to affect the progression, metastasis, and resistance to therapy depending on their state of reprogramming. Therapeutic benefit in targeting TAMs suggests that macrophages are attractive targets for cancer treatment. Chinese materia medica (CMM) is an important approach for treating cancer in China and in the Asian region. According to the theory of Chinese medicine (CM) and its practice, some prescriptions of CM regulate the body's internal environment possibly including the remodeling the tumor microenvironment (TME). Here we briefly summarize the pivotal effects of TAMs in shaping the TME and promoting tumorigenesis, invasion, metastasis and immunosuppression. Furthermore, we illustrate the effects and mechanisms of CMM targeting TAMs in antitumor therapy. Finally, we reveal the CMM's dual-regulatory and multi-targeting functions on regulating TAMs, and hopefully, provide the theoretical basis for CMM clinical practice related to cancer therapy

Extremely Low Genetic Diversity Indicating the Endangered Status of Ranodon sibiricus (Amphibia: Caudata) and Implications for Phylogeography

Background: The Siberian salamander (Ranodon sibiricus), distributed in geographically isolated areas of Central Asia, is an ideal alpine species for studies of conservation and phylogeography. However, there are few data regarding the genetic diversity in R. sibiricus populations. Methodology/Principal Findings: We used two genetic markers (mtDNA and microsatellites) to survey all six populations of R. sibiricus in China. Both of the markers revealed extreme genetic uniformity among these populations. There were only three haplotypes in the mtDNA, and the overall nucleotide diversity in the mtDNA was 0.00064, ranging from 0.00000 to 0.00091 for the six populations. Although we recovered 70 sequences containing microsatellite repeats, there were only two loci that displayed polymorphism. We used the approximate Bayesian computation (ABC) method to study the demographic history of the populations. This analysis suggested that the extant populations diverged from the ancestral population approximately 120 years ago and that the historical population size was much larger than the present population size; i.e., R. sibiricus has experienced dramatic population declines. Conclusion/Significance: Our findings suggest that the genetic diversity in the R. sibiricus populations is the lowest among all investigated amphibians. We conclude that the isolation of R. sibiricus populations occurred recently and was a result of recent human activity and/or climatic changes. The Pleistocene glaciation oscillations may have facilitated intraspecie