Nomograms incorporating hsa_circ_0029325 highly expressed in exosomes of hepatocellular carcinoma predict the postoperative outcomes.

BACKGROUND: Liquid biopsies, for example, exosomal circular RNA (circRNA) can be used to assess potential predictive markers for hepatocellular carcinoma (HCC) in patients after curative resection. This study aimed to search for effective prognostic biomarkers for HCC in patients after surgical resection based on exosomal circRNA expression profiles. We developed two nomograms incorporating circRNAs to predict the postoperative recurrence-free survival (RFS) and overall survival (OS) of HCC patients. METHOD: Plasma exosomes isolated from HCC patients and healthy individuals were used for circRNA microarray analysis to explore differentially expressed circRNAs. Pearson correlation analysis was used to evaluate the correlation between circRNAs and clinicopathological features. Cox regression analysis was used to explore the correlation between circRNA and postoperative survival time as well as recurrence time. A nomogram based on circRNA and clinicopathological characteristics was established and further evaluated to predict prognosis and recurrence. RESULT: Among 60 significantly upregulated circRNAs and 25 downregulated circRNAs, hsa_circ_0029325 was selected to verify its power for predicting HCC outcomes. The high expression level of exosomal hsa_circ_0029325 was significantly correlated with OS (P = 0.001, HR = 2.04, 95% CI 1.41-3.32) and RFS (P = 0.009, HR = 1.62, 95% CI 1.14-2.30). Among 273 HCC patients, multivariate regression analysis showed that hsa_circ_0029325 (HR = 1.96, 95% CI 1.21-3.18), tumor size (HR = 2.11, 95% CI 1.33-3.32), clinical staging (HR = 2.31, 95% CI 1.54-3.48), and tumor thrombus (HR = 1.74, 95% CI 1.12-2.7) were independent risk factors for poor prognosis in HCC patients after radical resection. These independent predictors of prognosis were incorporated into the two nomograms. The AUCs under the 1-year, 3-year, and 5-year survival and recurrence curves of the OS and RFS nomograms were 0.755, 0.749, and 0.742 and 0.702, 0.685, and 0.642, respectively. The C-index, calibration curves, and clinical decision curves showed that the two prediction models had good predictive performance. These results were verified in the validation cohort with 90 HCC patients. CONCLUSION: Our study established two reliable nomograms for predicting recurrence and prognosis in HCC patients. We also show that it is feasible to screen potential predictive markers for HCC after curative resection through exosomal circRNA expression profile analysis.

The immune response of hepatocellular carcinoma after locoregional and systemic therapies: The available combination option for immunotherapy.

Hepatocellular carcinoma (HCC) is associated with a highly heterogeneous immune environment that produces an immune response to various locoregional treatments (LRTs), which in turn affects the effectiveness of immunotherapy. Although LRTs still dominate HCC therapies, 50-60% of patients will ultimately be treated with systemic therapies and might receive those treatments for the rest of their life. TACE, SIRT, and thermal ablation can dramatically increase the immunosuppressive state of HCC, a condition that can be addressed by combination with immunotherapy to restore the activity of lymphocytes and the secretion of cellular immune factors. Immune treatment with locoregional and systemic treatments has dramatically changed the management of HCC. In this review, we examine the research on the changes in the immune microenvironment after locoregional or systemic treatment. We also summarize the regulation of various immune cells and immune factors in the tumor microenvironment and discuss the different infiltration degrees of immune cells and factors on the prognosis of HCC to better compare the efficacy between different treatment methods from the perspective of the tumor microenvironment. This information can be used to help develop treatment options for the upcoming new era of HCC treatment in the future.

Prolyl hydroxylase 2 inhibits glycolytic activity in colorectal cancer via the NF­κB signaling pathway.

A variety of malignancies preferentially meet energy demands through the glycolytic pathway. Hypoxia­induced cancer cell adaptations are essential for tumor development. However, in cancerous glycolysis, the functional importance and underlying molecular mechanism of prolyl hydroxylase domain protein 2 (PHD2) have not been fully elucidated. Gain­ and loss­of­function assays were conducted to evaluate PHD2 functions in colon cancer cells. Glucose uptake, lactate production and intracellular adenosine­5'­triphosphate/adenosine diphosphate ratio were measured to determine glycolytic activities. Protein and gene expression levels were measured by western blot analysis and reverse transcription­quantitative PCR, respectively. The human colon cancer xenograft model was used to confirm the role of PHD2 in tumor progression in vivo. Functionally, the data demonstrated that PHD2 knockdown leads to increased glycolysis, while PHD2 overexpression resulted in suppressed glycolysis in colorectal cancer cells. In addition, the glycolytic activity was enhanced without PHD2 and normalized after PHD2 reconstitution. PHD2 was shown to inhibit colorectal tumor growth, suppress cancer cell proliferation and improve tumor­bearing mice survival in vivo. Mechanically, it was found that PHD2 inhibits the expression of critical glycolytic enzymes (glucose transporter 1, hexokinase 2 and phosphoinositide­dependent protein kinase 1). In addition, PHD2 inhibited Ikkß­mediated NF­κB activation in a hypoxia­inducible factor­1α­independent manner. In conclusion, the data demonstrated that PHD2/Ikkß/NF­κB signaling has critical roles in regulating glycolysis and suggests that PHD2 potentially suppresses colorectal cancer.

Association between diet and the gut microbiome of young captive red-crowned cranes (Grus japonensis).

BACKGROUND: Exploring the association of diet and indoor and outdoor environments on the gut microbiome of red-crowned cranes. We investigated the microbiome profile of the 24 fecal samples collected from nine cranes from day 1 to 35. Differences in the gut microbiome composition were compared across diet and environments. RESULTS: A total of 2,883 operational taxonomic units (OTUs) were detected, with 438 species-specific OTUs and 106 OTUs common to the gut microbiomes of four groups. The abundance of Dietzia and Clostridium XI increased significantly when the red-crowned cranes were initially fed live mealworms. Skermanella and Deinococcus increased after the red-crowned cranes were fed fruits and vegetables and placed outdoors. Thirty-three level II pathway categories were predicted. Our study revealed the mechanism by which the gut microbiota of red-crowned cranes responds to dietary and environmental changes, laying a foundation for future breeding, nutritional and physiological studies of this species. CONCLUSIONS: The gut microbiome of red-crowned cranes could adapt to changes in diet and environment, but the proportion of live mealworms in captive red-crowned cranes can be appropriately reduced at the initial feeding stage, reducing the negative impact of high-protein and high-fat foods on the gut microbiome and growth and development.

Integrated proteogenomic characterization reveals an imbalanced hepatocellular carcinoma microenvironment after incomplete radiofrequency ablation.

BACKGROUND: Efforts to precisely assess tumor-specific T-cell immune responses still face major challenges, and the potential molecular mechanisms mediating hepatocellular carcinoma (HCC) microenvironment imbalance after incomplete radiofrequency ablation (iRFA) are unclear. This study aimed to provide further insight into the integrated transcriptomic and proteogenomic landscape and identify a new target involved in HCC progression following iRFA. METHODS: Peripheral blood and matched tissue samples were collected from 10 RFA-treated HCC patients. Multiplex immunostaining and flow cytometry were used to assess local and systemic immune responses. Differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) were explored via transcriptomic and proteogenomic analyses. Proteinase-3 (PRTN3) was identified in these analyses. And then, the ability of PRTN3 to predict overall survival (OS) was assessed in 70 HCC patients with early recurrence after RFA. In vitro CCK-8, wound healing and transwell assays were conducted to observe interactions between Kupffer cells (KCs) and HCC cells induced by PRTN3. The protein levels of multiple oncogenic factors and signaling pathway components were detected by western blotting. A xenograft mouse model was built to observe the tumorigenic effect of PRTN3 overexpression on HCC. RESULTS: Multiplex immunostaining revealed no immediate significant change in local immune cell counts in periablational tumor tissues after 30 min of iRFA. Flow cytometry showed significantly increased levels of CD4+ T cells, CD4+CD8+ T cells, and CD4+CD25+CD127- Tregs and significantly decreased the levels of CD16+CD56+ natural killer cells on day 5 after cRFA (p < 0.05). Transcriptomics and proteomics revealed 389 DEGs and 20 DEPs. Pathway analysis showed that the DEP-DEGs were mainly enriched in the immunoinflammatory response, cancer progression and metabolic processes. Among the DEP-DEGs, PRTN3 was persistently upregulated and closely associated with the OS of patients with early recurrent HCC following RFA. PRTN3 expressed in KCs may affect the migration and invasion of heat stress-treated HCC cells. PRTN3 promotes tumor growth via multiple oncogenic factors and the PI3K/AKT and P38/ERK signaling pathways. CONCLUSIONS: This study provides a comprehensive overview of the immune response and transcriptomic and proteogenomic landscapes of the HCC milieu induced by iRFA, revealing that PRTN3 promotes HCC progression after iRFA. TRIAL REGISTRATION: ChiCTR2200055606, http://www.chictr.org.cn/showproj.aspx?proj=32588 .

The combination of a prolonged treatment time window and alpha-fetoprotein benefits the tumor response of hepatocellular carcinoma patients as evaluated by the imRECIST: a single-center, retrospective study.

Background: The combined immunotargeting therapy of hepatocellular carcinoma (HCC) have brought remarkable results. There are still some drawbacks to the application of the immune-modified Response Evaluation Criteria in Solid Tumors to Immunotherapy (imRECIST). How many weeks does it take to confirm the true disease progression for HCC patients who had reported disease progression for the first time based on imRECIST. Whether alpha-fetoprotein (AFP), an important indicator in the progression and prognosis of liver cancer, has the same value in immunotherapy. This prompted more clinical data to gather evidence that the immunotherapy time window issue contradicts the potential benefit of therapy. Methods: This study retrospectively analyzed the clinical data of 32 patients who had undergone immunotherapy plus targeted therapy at the First Affiliated Hospital of Chongqing Medical University from June 2019 to June 2022. ImRECIST was used to evaluate the therapeutic efficacy among the patients. Before initial treatment and each immunotherapy cycle, each patient underwent standard abdominal computed tomography (CT) imaging and some biochemical indicators to assess physical condition and tumor response. All patients included will be divided into 8 groups. The differences in the survival outcomes of each treatment group were analysed. Results: Among the 32 advanced HCC patients, 9 patients achieved stable disease (SD), 12 patients showed progressive disease (PD), 3 patients showed a complete response (CR), and 8 patients showed a partial response (PR). There is no difference in baseline characteristics between subgroups. In relation to patients with PD, a prolonged therapeutic time window and the provision of continuous medication may lead to a PR, prolonging their overall survival (P=0.5864). Compared to the patients with continuous PD, there was no significant difference in the survival of patients with increased AFP concentrations after treatment who achieved PR or SD and ultimately showed PD (P=0.6600). Conclusions: In our study, the time window for treatment may need to be extended in the process of immunotherapy for HCC patients. An analysis of AFP may assist the imRECIST by providing a more accurate evaluation of tumor progression.

Unraveling the Emerging Niche Role of Hepatic Stellate Cell-derived Exosomes in Liver Diseases.

Hepatic stellate cells (HSCs) play an essential role in various liver diseases, and exosomes are critical mediators of intercellular communication in local and distant microenvironments. Cellular crosstalk between HSCs and surrounding multiple tissue-resident cells promotes or inhibits the activation of HSCs. Substantial evidence has revealed that HSC-derived exosomes are involved in the occurrence and development of liver diseases through the regulation of retinoid metabolism, lipid metabolism, glucose metabolism, protein metabolism, and mitochondrial metabolism. HSC-derived exosomes are underpinned by vehicle molecules, such as mRNAs and microRNAs, that function in, and significantly affect, the processes of various liver diseases, such as acute liver injury, alcoholic liver disease, nonalcoholic fatty liver disease, viral hepatitis, fibrosis, and cancer. As such, numerous exosomes derived from HSCs or HSC-associated exosomes have attracted attention because of their biological roles and translational applications as potential targets for therapeutic targets. Herein, we review the pathophysiological and metabolic processes associated with HSC-derived exosomes, their roles in various liver diseases and their potential clinical application.

Sublethal chlorantraniliprole exposure induces autophagy and apoptosis through disrupting calcium homeostasis in the silkworm Bombyx mori.

The intensive application of chlorantraniliprole (CAP) leaves residues in the environment, posing a potential threat to non-target organisms. In the present study, we investigated the adverse effects of sublethal CAP exposure on Bombyx mori. Sublethal CAP (0.02 mg/L) was shown to induce the release of intracellular Ca2+ in BmN cells. Meanwhile, Ca2+ -dependent genes were induced in the midgut at 72 h after CAP (0.01 mg/L) exposure, and damaged mitochondria, autophagosomes, nuclear membrane rupture and condensed chromatin were observed. Moreover, the key genes in the oxidative phosphorylation pathway were significantly down-regulated. The transcript levels of autophagy-related genes ATG6 and ATG8 were significantly up-regulated, and the protein levels of LC3-II and ATG7 were significantly increased by 3.72- and 3.33-fold, respectively. Additionally, the transcript levels of the upstream genes in the apoptosis pathway (calpain and Apaf-1) were significantly up-regulated, the protein levels of the downstream gene caspase 3 and its cleaved form were significantly up-regulated by 1.97- and 4.55-fold, respectively, consistent with the elevated caspase 3 activity at 72 h. Collectively, these findings demonstrate that intracellular Ca2+ release induced by sublethal CAP inhibits oxidative phosphorylation pathway, which causes mitochondrial dysfunction, leading to autophagy and apoptosis in the midgut of B. mori.

Mechanism of autophagy induced by low concentrations of chlorantraniliprole in silk gland, Bombyx mori.

Chlorantraniliprole (CAP) is widely used in the control of agricultural pests, and its residues can affect the formation of silkworm (Bombyx. mori) cocoon easily. To accurately evaluate the toxicity of CAP to silkworms and clarify the mechanism of its effect on silk gland function, we proposed a novel toxicity evaluation method based on the body weight changes after CAP exposure. We also analyzed the Ca2+-related ATPase activity, characterized energy metabolism and transcriptional changes about the autophagy key genes on the downstream signaling pathways. The results showed that after a low concentration of CAP exposed for 96 h, there were CAP residues in the silk glands of B. mori, the activities of Ca2+-ATPase and Ca2+-Mg2+-ATPase decreased significantly (P ≤ 0.01), and the activation of AMPK-related genes AMPK-α and AMPK-ß were up-regulated by 6.39 ± 0.02-fold and 12.33 ± 1.06-fold, respectively, reaching a significant level (P ≤ 0.01)). In addition, the autophagy-related genes Atg1, Atg6, Atg5, Atg7, and Atg8 downstream AMPK were significantly up-regulated at 96 h (P ≤ 0.05). The results of immunohistochemistry and protein expression assay for autophagy marker Atg8 further confirmed the occurrence of autophagy. Overall, our results indicate that CAP exposure leads to autophagy in the silk gland of B. mori and affects their physiological functions, which provides guidance for the evaluation of toxicity of low concentration environmental CAP residues to insects.

A selection marker-free method for gene deletion and editing in baculovirus genomes.

Here, we develop a simple, efficient, bacmid-based, selection marker-free method for gene deletion and editing in baculovirus genomes. Specifically, based on pFastbac1, a donor plasmid with long left and right homology arms but without a reporter was constructed for disrupting ie1, an essential baculovirus gene. Instead of ligating with a plasmid, the homology arms were introduced to the polyhedrin locus of BmNPV bacmid using the BmNPV bac-to-bac expression system. Two viruses generated from the modified bacmid and unmodified BmNPV bacmid were then used to co-infect BmN cells in order that recombination takes place at the ie1 locus between them. Finally, without multiple rounds of purification, total cellular DNA was isolated, transformed into Cacl2-treated competent DH10B cells, and then blue colonies were selected for PCR screening. Remarkably, the proportion of blue colonies containing ie1-disrupted bacmid was found to be around 7 %. Moreover, using primers flanking the homology arms further confirmed that all these positive recombinants were double crossovers. These findings indicate that our method is also capable of gene modification if inverse PCR or seamless cloning is used to construct the donor plasmid and sequencing is employed to select positive colonies.

A well-matched marriage of immunotherapy and radiofrequency ablation to reduce the relapse and progression of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) remains a health challenge with increasing incidence worldwide. Radiofrequency ablation (RFA) is a potentially curative option for patients with early-stage HCC. However, the high rate of tumor recurrence limits long-term survival when the tumors are larger than 2 cm and undergoing insufficient RFA (iRFA). Notably, in situ tumor necrosis due to thermal ablation is assumed to be a source of antigens that induce antitumor immunity. Therefore, mounting studies and trials have attempted to provide a rational and effective therapeutic strategy combining RFA and immunotherapy to treat HCC. Nowadays, many controversies and challenges with this combined therapeutic strategy remain to be resolved, such as the indications for adjuvant immunotherapy along with RFA in early HCC, the sequence of the two treatments in advanced HCC, and the optimal timing of immunotherapy before or after RFA. In addition, individualized treatment strategies need to be perfected for patients with HCC.

Hepatic stellate cell exosome-derived circWDR25 promotes the progression of hepatocellular carcinoma via the miRNA-4474-3P-ALOX-15 and EMT axes.

Recently, the emerging role of circular RNAs (circRNAs) in tumor development and progression has been a topic of great interest. Nevertheless, the effects of hepatic stellate cell (HSC)-derived exosomes in hepatocellular carcinoma (HCC) remain unclear. Here, we aim to explore the potential effect of HSC exosome-derived circWDR25 on the aggressiveness of HCC. Firstly, a microarray analysis of circRNAs was performed to profile and identify the differentially expressed circRNAs derived from HSC exosomes activated by HCC cells. Subsequently, the roles of circWDR25 in HCC tumor growth and aggressiveness were confirmed through in vitro and in vivo functional experiments. Moreover, RNA pull-down, dual-luciferase reporter assays, and fluorescent in situ hybridization (FISH) were performed to determine interactions in the circWDR25-miR-4474-3p-ALOX15 loop. Immunohistochemical analysis was also performed on a microarray of HCC tissues and peritumoral tissues. We found that overexpressed peritumoral circWDR25 was associated with survival and recurrence in patients with HCC and promoted the progression of HCC cells both in vitro and in vivo. Mechanistically, both exogenous and HSC exosomal-derived circWDR25 regulated the expression of ALOX15 by sponging miR-4474-3p and ultimately inducing an epithelial-to-mesenchymal transition (EMT) in HCC cells. Moreover, exogenous and HSC exosomal-derived circWDR25 promoted the expression of CTLA-4 in HSCs and PD-L1 in HCC cells. In conclusion, circWDR25 facilitated HCC cell proliferation and invasion via the circWDR25/miR-4474-3p/ALOX15 and EMT axes and it promoted the expression of CTLA-4 in HSCs and PD-L1 in HCC cells, thus providing insights into the mechanism of tumor aggressiveness mediated by HSC-derived exosomal circWDR25.

Changes in the Diversity and Composition of Gut Microbiota of Red-Crowned Cranes (Grus japonensis) after Avian Influenza Vaccine and Anthelmintic Treatment.

Gut microbiota homeostasis is important for host health and well-being; however, drugs may affect the composition and function of the gut microbiota. Red-crowned cranes are a vulnerable species. Treatment of red-crowned cranes with avian influenza vaccines and anthelmintics has played pivotal roles in therapeutic management in zoos. To investigate the changes in the diversity and composition of gut microbiota after the avian influenza vaccine and anthelmintic treatment, we used 16S rRNA sequencing to obtain and compare the bacterial community composition before and after the treatment. The alpha diversity of the gut microbiota of red-crowned cranes decreased on the day of the treatment and then fluctuated over time. The composition of gut microbiota tended to be similar in the short term after the treatment, as supported by the beta diversity hierarchical cluster analysis. Only 3, 8, and 72 operational taxonomic units (OTUs) of the three individuals were shared among the five groups before and after treatment. The relative abundance of Firmicutes significantly increased to 99.04% ± 0.28% on the day of the treatment, in which the relative abundance of Lactobacillus was 93.33% ± 5.85%. KEGG pathways analysis indicated that the main function of the gut microbiota is involved in metabolism, and the present study indicates that the gut microbiota of red-crowned cranes is resilient to the avian influenza vaccine and anthelmintic, even disordered in the short term, and could recover over time. More individual experimentation and functional potential in metabolism are needed in the future to support animal disease control and optimal management in the zoo.

Effect of Tachinid Parasitoid Exorista japonica on the Larval Development and Pupation of the Host Silkworm Bombyx mori.

The Tachinidae are natural enemies of many lepidopteran and coleopteran pests of crops, forests, and fruits. However, host-tachinid parasitoid interactions have been largely unexplored. In this study, we investigated the effects of tachinids on host biological traits, using Exorista japonica, a generalist parasitoid, and the silkworm Bombyx mori, its lepidopteran host, as models. We observed that E. japonica parasitoidism did not affect silkworm larval body weight gain and cocooning rate, whereas they caused shortened duration of molting from the final instar to the pupal stage, abnormal molting from larval to pupal stages, and a subsequent decrease in host emergence rate. Moreover, a decrease in juvenile hormone (JH) titer and an increase in 20-hydroxyecdysone (20E) titer in the hemolymph of parasitized silkworms occurred. The transcription of JH and 20E responsive genes was downregulated in mature parasitized hosts, but upregulated in parasitized prepupae while Fushi tarazu factor 1 (Ftz-f1), a nuclear receptor essential in larval ecdysis, showed dramatically reduced expression in parasitized hosts at both the mature and prepupal stages. Moreover, the transcriptional levels of BmFtz-f1 and its downstream target genes encoding cuticle proteins were downregulated in epidermis of parasitized hosts. Meanwhile, the content of trehalose was decreased in the hemolymph, while chitin content in the epidermis was increased in parasitized silkworm prepupae. These data reveal that the host may fine-tune JH and 20E synthesis to shorten developmental duration to combat established E. japonica infestation, while E. japonica silences BmFtz-f1 transcription to inhibit host pupation. This discovery highlights the novel target mechanism of tachinid parasitoids and provides new clues to host/tachinid parasitoid relationships.

Evaluation of tolerance to λ-cyhalothrin and response of detoxification enzymes in silkworms reared on artificial diet.

A representative silkworm rearing mode of Ⅰ-Ⅲ instars reared on artificial diet and Ⅳ-Ⅴ instars reared on fresh mulberry leaves has been recognized in some sericultural areas of China. Under this rearing mode, silkworms are prone to be poisoned by pesticide residues on mulberry leaves at the Ⅳ and Ⅴ instar stages. As one of the most widely applied insecticides, λ-cyhalothrin was used to study the insecticide tolerance of silkworm reared on artificial diet (referred as the AD group). Our results showed that the newly ecdysized Ⅳ instar larvae in the AD group were less tolerant to λ-cyhalothrin compared to the mulberry leaves reared group (referred as the ML group). After continuous exposure to trace λ-cyhalothrin, the weight gain and the survival rate of silkworms were significantly lower in the AD group than those in the ML group, even though compensatory growth was observed in the control of the AD group. Histopathology and ultrastructure of fat body showed that λ-cyhalothrin induced more severe cell injuries in the AD group, such as shrunken nucleus, dilatation of endoplasmic reticulum, and mitochondrial swelling. The transcription levels of detoxification related genes (CYP4M5, CYP6AB4, CarE2, CarE5, GSTe1 and GSTe3) and the enzyme activities of P450s, CarEs and GSTs were inducible by trace λ-cyhalothrin in a time-specific manner, and the data showed that the response of P450 enzyme activity was retarded in the AD group, indicating a potential reason for a higher sensitivity to λ-cyhalothrin. Our results provided a new clue for the study of the relationship between feed nutrition and detoxification ability, and also provided an important reference for the development of modern silkworm rearing mode.

Damaging effects of TiO2 nanoparticles on the ovarian cells of Bombyx mori.

As a new type of biologically compatible material, TiO2 NPs are widely used in the industry as additives, drug carriers, and components of skin care products. Due to their wide use, residual TiO2 NPs in the environment are a safety concern that has attracted extensive attention. In this study, the ovarian cell line BmN of the model organism Bombyx mori was used to reveal the damaging effects of TiO2 NPs exposure. The results demonstrated that TiO2 NPs exhibited a dose-dependent effect on the relative cell viability, with significant toxic effects being observed above 20 mg/L. Oxidative damage analysis showed that ROS accumulated significantly in BmN cells after exposure to TiO2 NPs (P ≤ 0.05) and induced DNA damage. Further analysis revealed that the transcriptional levels of key superoxide dismutase genes (SOD) decreased significantly, while the transcriptions of key genes of the MAPK/NF-κB signaling pathway (P38, MEK, ERK and REL) and the downstream inflammatory factor genes (IL6 and TNFSF5) were all significantly up-regulated (P ≤ 0.05). Overall, our results indicate that exposure to TiO2 NPs leads to reduced transcription of antioxidant genes, accumulation of peroxides, and inflammation. These findings provide valuable data for the safety evaluation of environmental residues of TiO2 NPs.

Comprehensive transcriptome characterization of Grus japonensis using PacBio SMRT and Illumina sequencing.

The red-crowned crane (Grus japonensis) is an endangered species distributed across southeast Russia, northeast China, Korea, and Japan. Here, we sequenced for the first time the full-length unreferenced transcriptome of red-crowned crane mixed samples using a PacBio Sequel platform. A total of 359,136 circular consensus sequences (CCS) were obtained via clustering to remove redundancy. A total of 303,544 full-length non-chimeric sequences were identified by judging whether CCS contained 5' and 3' adapters, and the poly(A) tail. Eight samples were sequenced using Illumina, and PacBio sequencing data were corrected according to the collected Illumina data to obtain more accurate full-length transcripts. A total of 4,100 long non-coding RNAs, 13,115 simple sequences repeat loci and 29 transcription factor families were identified. The expression of lncRNAs and TFs in pancreas was lowest comparing with other tissues. Many enriched immune-related transmission pathways (MHC and IL receptors) were identified in the spleen. This study will contribute to a better understanding of the gene structure and post-transcriptional regulatory network, and provide references for future studies on red-crowned cranes.

Imbalance of intestinal microbial homeostasis caused by acetamiprid is detrimental to resistance to pathogenic bacteria in Bombyx mori.

The neonicotinoid insecticide acetamiprid is widely applied for pest control in agriculture production, and its exposure often results in adverse effects on a non-target insect, Bombyx mori. However, only few studies have investigated the effects of exposure to sublethal doses of neonicotinoid insecticides on gut microbiota and susceptibility to pathogenic bacteria. In this study, we aimed to explore the possible mechanisms underlying the acetamiprid-induced compositional changes in gut microbiota of silkworm and reduced host resistance against detrimental microbes. This study indicated that sublethal dose of acetamiprid activated the dual oxidase-reactive oxygen species (Duox-ROS) system and induced ROS accumulation, leading to dysregulation of intestinal immune signaling pathways. The evenness and structure of bacterial community were altered. Moreover, after 96 h of exposure to sublethal dose of acetamiprid, several bacteria, such as Pseudomonas sp (Biotype A, DOP-1a, XW34) and Staphylococcus sp (RCB1054, RCB314, X302), invaded the silkworm hemolymph. The survival rate and bodyweight of the acetamiprid treated silkworm larvae inoculated with Enterobacter cloacae (E. cloacae) were significantly lower than the acetamiprid treatment group, suggesting that acetamiprid reduced silkworm resistance against pathogens. These findings indicated that acetamiprid disturbed gut microbial homeostasis of Bombyx mori, resulting in changes in gut microbial community and susceptibility to detrimental microbes.

The mitochondrial genome and phylogenetic characteristics of the Thick-billed Green-Pigeon, Treron curvirostra: the first sequence for the genus.

Members of the genus Treron (Columbidae) are widely distributed in southern Asia and the Indo-Malayan Region but their relationships are poorly understood. Better knowledge of the systematic status of this genus may help studies of historical biogeography and taxonomy. The complete mitochondrial genome of T. curvirostra was characterized, a first for the genus. It is 17,414 base pairs in length, containing two rRNAs, 22 tRNAs, 13 protein coding genes (PCGs), and one D-loop with a primary structure that is similar to that found in most members of Columbidae. Most PCGs start with the common ATG codon but are terminated by different codons. The highest value of the Ka/Ks ratio within 13 PCGs was found in ATP8 with 0.1937, suggesting that PCGs of the mitochondrial genome tend to be conservative in Columbidae. Moreover, the phylogenetic relationships within Columbidae, which was based on sequences of 13 PCGs, showed that (T. curvirostra + Hemiphaga novaeseelandiae) were clustered in one clade, suggesting a potentially close relationship between Treron and Hemiphaga. However, the monophyly of the subfamilies of Columbidae recognized by the Interagency Taxonomic Information System could not be corroborated. Hence, the position of the genus Treron in the classification of Columbidae may have to be revised.

Cloning and functional analysis of autophagy-related gene 7 in Bombyx mori, silkworm.

Silkworm (Bombyx mori) is an important economic insect and an attractive model system. A series of autophagy-related genes (Atgs) are involved in the autophagic process, and these Atgs have been proved to play important roles in the development. Atg7 stands at the hub of two ubiquitin-like systems involving Atg8 and Atg12 in the autophagic vesicle. In the present study, we cloned and characterized a BmAtg7 gene in Bombyx mori. The open reading frame (ORF) of BmAtg7 was 1908 bp in length, and it encoded a polypeptide of 635 amino acids. BmAtg7 was highly expressed in the posterior silk gland, fatbody, and epidermis. The expression profile of BmAtg7 in the fatbody showed an increasing tendency from day 1 of the 5th instar to the prepupal stage. After chlorantraniliprole (CAP) exposure, the transcriptional level of BmAtg7 was continuously decreased. After depletion of BmAtg7 by RNAi, the expressions of BmAtg7, BmAtg8, and BmEcr were all downregulated, while the expression of BmJHBP2 was upregulated. However, depletion of BmAtg7 did not prevent the metamorphosis of silkworm from larvae to pupae, while the occurrence of such process was delayed. After the 20-hydroxyecdysone (20E) treatment, the expression characteristics of these four genes (BmAtg7, BmAtg8, BmEcr and BmJHBP2) were contrary to the results after depletion of BmAtg7. Our results suggested that although CAP exposure could significantly inhibit the expression of BmAtg7 continuously, the changes of BmAtg7 was not the key factor in CAP-induced metamorphosis defects.