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Identifying populations that benefit from immune checkpoint blockade (ICB) therapy remains a major challenge in the treatment of lung adenocarcinoma (LUAD). Existing programmed cell death (PCD) related prognostic models only consider a single mechanism, such as ferroptosis, necroptosis, and pyroptosis, and do not reflect the interaction of multiple mechanisms. This study aims to explore lncRNAs associated with multiple modes of PCD and reveal a risk signature to assess prognosis and treatment outcomes in LUAD patients.Based on expression data in The Cancer Genome Atlas (TCGA) database, ferroptosis, necroptosis, and pyroptosis-related lncRNAs (FNPRlncRNAs) were obtained by taking the intersection of ferroptosis-related lncRNAs (FRlncRNAs), necroptosis-related lncRNAs (NRlncRNAs), and pyroptosis-related lncRNAs (PRlncRNAs) differentially expressed in LUAD and normal tissues. Patients with complete survival information and expression data from TCGA database were randomly assigned to training and testing sets (1:1). Univariate, LASSO, and multivariate Cox regression analyses were performed on the training set, and a risk signature was established. Kaplan-Meier survival curves were used to verify the prognostic ability of risk signature, and receiver operating characteristic (ROC) curves were used to assess the predictive accuracy. We then analyzed molecular and immune profile differences between high and low-risk subgroups. T-cell dysfunction and Exclusion (TIDE) scores were used to assess the response to immunotherapy in each risk subgroup. Finally, three LUAD clusters (C1, C2, and C3) were identified according to the risk signature.Patients in the low-risk subgroup had higher overall survival (OS) than that in the high-risk subgroup in the K-M survival curve. The area under ROC curves (AUC) of 1-, 3-, and 5-year ROC were 0.742, 0.762, and 0.749 in the training set, and 0.672, 0.642, and 0.563 in the testing set, respectively. Compared with the high-risk subgroup, patients in the low-risk subgroup have beneficial tumor immune microenvironment and molecular characteristics, but are less likely to benefit from immunotherapy. Finally, the three LUAD clusters (C1, C2, C3) identified by risk signature had different responses to drug treatment.The prognosis risk signature constructed using FNPRlncRNAs is helpful to predict the prognosis of LUAD and may contribute to its individualized treatment.

Background Most human diseases are accompanied by systems changes. Systems biomarkers should reflect such changes. The phosphorylation and dephosphorylation of biomolecules maintain human homeostasis. However, the systems biomarker characteristics of circulating alkaline phosphatase, a routine blood test conducted for many human diseases, have never been investigated. Method This study retrieved the circulating alkaline phosphatase (ALP) activities from patients with 48 clinically confirmed diseases and healthy individuals from the database of our hospital during the past five years. A detailed analysis of the statistical characteristics of ALP was conducted, including quantiles, receiving operator curve (ROC), and principal component analysis. Results Among the 48 diseases, 45 had increased, and three had decreased median levels of ALP activities compared to the healthy control. Preeclampsia, hepatic encephalopathy, pancreatic cancer, and liver cancer had the highest median values, whereas nephrotic syndrome, lupus erythematosus, and nephritis had decreased median values compared to the healthy control. Further, area under curve (AUC) values were ranged between 0.61-0.87 for 19 diseases, and the ALP activities were the best systems biomarker for preeclampsia (AUC 0.87), hepatic encephalopathy (AUC 0.87), liver cancer (AUC 0.81), and pancreatic cancer (AUC 0.81). Conclusions Alkaline phosphatase was a decent systems biomarker for 19 different types of human diseases. Understanding the molecular mechanisms of over-up-and-down-regulation of ALP activities might be the key to understanding the whole-body systems' reactions during specific disease progression.

Land use function transition can change the eco-environment. To achieve an “Intensive and efficient production space, moderately livable living space, and beautiful ecological space”, the ecological effects of land use function transition in the context of ecologically fragile areas and rapidly developing areas of socio-economic importance need to be studied. In this study, from the perspective of “production-living-ecological” spaces, we calculated the index of regional eco-environment quality, positive and negative effects of eco-environment impact, and the ecological contribution rate and analyzed the driving factors. We found the following: (1) The production space was greatly compressed, living space was expanded, and ecological space was significantly squeezed. Haikou underwent a rapid transformation from an agriculture-dependent city to an industrial city. Land supply for urban and rural living was guaranteed by the Chinese land management department. However, Haikou prioritized economic development over environmental protection. (2) The regional eco-environment quality index decreased from 2009 to 2018. The expansion of pasture-based ecological spaces is important for improving the quality of the eco-environment, and the reduction of forest ecological space strongly influences the deterioration of the eco-environment. (3) Resource base, historical level of utilization, suitability of land, the ecological value potentiality, and regional policies greatly affected land use function transition and its eco-environment. (4) Refining the planning ofterritorial space, comprehensively improving land and resources, and reforming the rural land system greatly influenced policy guidance and technical regulation for coordinating “production-living-ecological” spaces and improving the regional eco-environment. In this study, we tested the effect of regional policy regulation on land use function transition and provided a reference for coordinating “production-living-ecological” spaces.

Land use function transition can change the eco-environment. To achieve an “Intensive and efficient production space, moderately livable living space, and beautiful ecological space”, the ecological effects of land use function transition in the context of ecologically fragile areas and rapidly developing areas of socio-economic importance need to be studied. In this study, from the perspective of “production-living-ecological” spaces, we calculated the index of regional eco-environment quality, positive and negative effects of eco-environment impact, and the ecological contribution rate and analyzed the driving factors. We found the following: (1) The production space was greatly compressed, living space was expanded, and ecological space was significantly squeezed. Haikou underwent a rapid transformation from an agriculture-dependent city to an industrial city. Land supply for urban and rural living was guaranteed by the Chinese land management department. However, Haikou prioritized economic development over environmental protection. (2) The regional eco-environment quality index decreased from 2009 to 2018. The expansion of pasture-based ecological spaces is important for improving the quality of the eco-environment, and the reduction of forest ecological space strongly influences the deterioration of the eco-environment. (3) Resource base, historical level of utilization, suitability of land, the ecological value potentiality, and regional policies greatly affected land use function transition and its eco-environment. (4) Refining the planning of territorial space, comprehensively improving land and resources, and reforming the rural land system greatly influenced policy guidance and technical regulation for coordinating “production-living-ecological” spaces and improving the regional eco-environment. In this study, we tested the effect of regional policy regulation on land use function transition and provided a reference for coordinating “production-living-ecological” spaces.

This paper was aimed at providing a perspective on the assessment of land-use dominant transition. Based on the transfer matrix of land-use type, the dynamic degree of dominant morphology transition was measured, and an active degree index was proposed. The spatiotemporal differentiation characteristics were assessed and analyzed by land use change characteristics in different phases. Our assessment resulted in three types of spatiotemporal differentiation of Hainan Province in China. The whole island can be divided into three regions with two parallel lines at 45° based on spatiotemporal differentiation characteristics. The "slow-type" was distributed in four eastern counties (cities), "steady-type" was distributed in 13 central and western counties (cities), and the "active-type" was distributed in four central counties (cities). Over three phases, namely 2010-2011, 2012-2015, and 2015-2018, four levels of spatiotemporal differentiation of the 21 counties (cities) were assessed, and they were relatively stable, leaping, declining, and unstable. Areas of new construction and fixed assets investments consumed by increased construction land were the positive factors of land-use dominant transition, while output values of secondary industries and area of industrial-mining per capita were the negative factors. Based on these results, a more informative examination of LULC was proposed, and all resulting land management policies will be more targeted and effective.

Background Lung adenocarcinoma (LUAD), which is the most common subtype of non-small cell lung cancer, is a leading course of cancer-related mortality worldwide. Recently, circular RNA (CircRNAs) has become a hot spot in cancer research because of its important role in tumorigenesis and development and its superior stability. This study aims to clarify the role of circ-AASDH in LUAD and explore its competitive endogenous RNA mechanism. Methods The circ-AASDH, miR-140-3p and E2F transcription factor 7 (E2F7) mRNA expression levels were detected via qRT-PCR. CCK-8 and colony formation assay were used to evaluate the ability of cell proliferation. Transwell assay and wound healing assay were performed to measure the invasion and migration ability. Flow cytometry was used to detect the apoptosis of cells. Moreover, Sanger sequencing, RNaseR treatment and divergent primers were used to verify the circular structure. Luciferase reporter and RNA pull-down experiment were performed to characterize the ceRNA mechanism of circ-AASDH. The xenograft model of mice was established to investigate the tumorigenicity of circ-AASDH to LUAD in vivo. Results By screening for differentially expressed circRNAs, we found that circ-AASDH was highly expressed in LUAD tissues and cells and correlated with tumor size, clinical stage and poor prognosis. Transfection of si-circ-AASDH can inhibit the proliferation and migration of LUAD cells and promote apoptosis in vitro. In mechanism, circ-AASDH could be used as a sponge of miR-140-3p to weaken its inhibition on the expression of E2F7. Additionally, the overexpression of circ-AASDH could deduce the suppression of miR-140-3p on the malignant progression of LUAD cells. Besides, silencing of circ-AASDH inhibited cell proliferation and migration by regulating the expression of E2F7. Furthermore, overexpression of circ-AASDH can promote the growth of LUAD in vivo. Conclusions Circ-AASDH/miR-140-3p/E2F7 regulating axis promoted the progression in LUAD. Our results provided ideas for understanding the biological mechanism of circ-AASDH and clarify potential therapeutic targets in LUAD.

Gefitinib, the first-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), has become the standard of care for the first-line of therapy for advanced non-small cell lung cancer (NSCLC) with common EGFR mutation. However, the efficacy of preoperative gefitinib therapy in patients with common EGFR mutations remains poorly defined. We describe a NSCLC patient with bilateral synchronous lesions who had a significantly positive response to gefitinib before radical surgical resection. At the time of initial diagnosis, we were unable to confirm whether the two lesions were metastatic or synchronous primary lesions. Accordingly, we performed CT-guided percutaneous left lung biopsy resulting in a diagnosis of lung adenocarcinoma with exon 21 L858R point mutation of EGFR, This diagnosis was followed by preoperative gefitinib therapy for 8 weeks leading to a significant reduction in the lesion in the left lower lobe. Then the left lower lobectomy and mediastinal lymphadenectomy were performed. In addition, 3 months following resection of the left lower lobe tumor the patient underwent a right lower lobe wedge resection. This report indicates that NSCLC patient harboring common EGFR mutation accepting the first-generation EGFR-TKI gefitinib as a neoadjuvant targeted therapy option is safe, feasible, and well-tolerated.

BACKGROUND Affected by the COVID-19, a great many patients were quarantined and could not go to the hospital, which made the outpatient follow-up unable to be carried out normally. OBJECTIVE Our study aimed to assess the feasibility of a postoperative follow-up cloud platform based on a standardized discourse form among postoperative patients with lung cancer in COVID-19 pandemic. METHODS The postoperative follow-up cloud platform was established by designing a standardized discourse form, and constructing a follow-up database and a health education center. The follow-up team was comprised of fixed and experienced full-time doctors. A total of 184 and 178 patients as the intervention and control group, who were followed up by the cloud platform and outpatient department. At the end of the follow-up period, our center carried out the satisfaction with follow-up and SF-36 health questionnaire survey. RESULTS There was no statistically significant difference in the baseline characteristics of the patients between the two groups. At the end of the follow-up period, satisfaction with follow-up in the intervention group was better than that in the control group (96.74%, 178/184 vs 87.08%, 155/178; P＜0.001). The intervention group had the similar results compared with the control group in SF-36 scales (Average, 64.79±13.59 vs 63.07±15.00; P=0.256). CONCLUSIONS The postoperative follow-up cloud platform based on a standardized discourse form can break the limitations of time and space, improve medical service and doctor-patient communication, and promote scientific research, which is feasible in COVID-19 pandemic.

Visceral pleural invasion (VPI) has been identified as an adverse prognostic factor for non‐small cell lung cancer (NSCLC). Accurate nodal staging for NSCLC correlates with improved survival, but it is unclear whether tumors with VPI require a more extensive lymph nodes (LNs) dissection to optimize survival. We aimed to evaluate the impact of VPI status on the optimal extent of LNs dissection in stage I NSCLC, using the Surveillance, Epidemiology, and End Results (SEER) database. We identified 9297 surgically treated T1‐2aN0M0 NSCLC patients with at least one examined LNs. Propensity score matching was conducted to balance the baseline clinicopathologic characteristics between the VPI group and non‐VPI group. Log‐rank tests along with Cox proportional hazards regression methods were performed to evaluate the impact of extent of LNs dissection on survival. VPI was correlated with a significant worse survival, but there was no significant difference in survival rate between PL1 and PL2. Patients who underwent sublobectomy had slightly decreased survival than those who underwent lobectomy. Pathologic LNs examination was significantly correlated with survival. Examination of 7‐8 LNs and 14‐16 LNs conferred the lowest hazard ratio for T1‐sized/non‐VPI tumors (stage IA) and T1‐sized/VPI tumors (stage IB), respectively. The optimal extent of LNs dissection varied by VPI status, with T1‐sized/VPI tumors (stage IB) requiring a more extensive LNs dissection than T1‐sized/non‐VPI tumors (stage IA). These results might provide guidelines for surgical procedure in early stage NSCLC.

BACKGROUND: We sought to determine the perioperative safety and feasibility outcomes of stage IIIA (N2) non-small cell lung cancer (NSCLC) following neoadjuvant immunotherapy or neoadjuvant chemotherapy. METHODS: The clinical details of patients who attended the Affiliated Hospital of Qingdao University between January 2019 and December 2020 were retrospectively evaluated. Eligible patients had pathologically proven stage IIIA (N2) NSCLC and were randomly prescribed neoadjuvant therapy. Those in the neoadjuvant immunotherapy group received two cycles of nivolumab (3 mg/kg) and those in the control group received neoadjuvant chemotherapy (1,000 mg/m(2) gemcitabine and 80 mg/m(2) cisplatin). All patients were scheduled to undergo surgery. The primary endpoint was the risk of major complications within 30 days of surgery and the secondary endpoints were interval to surgery and 30-day mortality. RESULTS: A total of 107 eligible patients were evaluated of whom 25 were allocated to the neoadjuvant immunotherapy group and 82 to the neoadjuvant chemotherapy group. The median interval to surgery was similar in the two groups at 29.2 days [95% confidence interval (CI), 27.1 to 31.4 days] in the immunotherapy group and 28.7 days (95% CI, 27.6 to 29.8 days) in the chemotherapy group (P=0.656). While treatment-related adverse events were reported in most patients, all 25 patients completed two cycles of neoadjuvant immunotherapy and 80 of 82 patients completed two cycles of neoadjuvant chemotherapy, although one patient in the latter group died within 30 days of surgery. There was no statistically significant difference between the groups in the probability of grade 3 or higher postoperative complications within 30 days after surgery (P=0.757). CONCLUSIONS: Most patients achieved the primary and secondary endpoints of the study. However, the major pathological response (MPR) showed statistically significant differences between the neoadjuvant immunotherapy and neoadjuvant chemotherapy groups.

Background As a type of non-coding RNA, circular RNAs (circRNAs) are considered to be functional molecules associated with human cancers. An increasing number of circRNAs have been verified in malignant progression in a number of cancers. The circRNA, circFBXW7, has been proven to play an important role in tumor proliferation and metastasis. However, whether circFBXW7 influences progression in lung adenocarcinoma (LUAD) remains unclear. Methods Quantitative real-time reverse transcriptase PCR (qRT-PCR) was used to verify circFBXW7 in LUAD cell lines and LUAD tissues. Kaplan-Meier analysis was then used to compare the disease-free survival (DFS) and overall survival (OS) of these LUAD patients. The biological function of circFBXW7 was examined by overexpression and knockdown of circFBXW7 using MTT assay, EdU assay, wound-healing assay, and Transwell in vitro assays. To explore the mechanism of the circFBXW7, RNA pull-down assay, dual luciferase reporter assay, and RNA immunoprecipitation (RIP) assay were employed to examine the interaction between circFBXW7 and miR-942-5p. Western blot was used to study the fundamental proteins associated with the epithelial-mesenchymal transition (EMT) pathway. In vivo studies with BALB/c nude mice subcutaneously injected with cells stably overexpressing circFBXW7 were performed to further validate the in vitro results. Results circFBXW7 was downregulated in LUAD cell lines and tissues, and LUAD patients with lower levels had shorter DFS and OS. The in vitro study showed that circFBXW7 overexpression inhibited proliferation and migration of A549 and HCC2279 cell lines. These results were confirmed by circFBXW7 knockdown, which showed the reverse effect. The in vivo model showed that the circRNA levels influenced the tumor growth. Finally, we determined that circFBXW7 target miRNA-942-5p which regulates the EMT gene BARX2. The modulation of circFBXW7 levels produced significant changes in EMT genes in vitro and in vivo. Conclusions Our findings showed that circFBXW7 inhibits proliferation and migration by controlling the miR-942-5p/BARX2 axis in LUAD cell lines and its levels correlates with patient survival suggesting that regulating circFBXW7 could have therapeutic value in treating LUAD patients.

Testosterone (T) is essential for muscle fiber formation and growth. However, the specific mechanism by which T regulates skeletal muscle development in chicken embryos remains unclear. In this study, the role of T in myoblast proliferation both in vivo and in vitro was investigated. Results showed that the T administration significantly increased the ratio of breast muscle and leg muscle. T induced a significant increase in the cross-sectional area (CSA) and density of myofiber and the ratio of PAX7-positive cells in the skeletal muscle. Exogenous T also induced the upregulation of myogenic regulatory factors (MRFs) and cyclin-dependent kinases (CDK2)/Cyclin D1 (CCND1) and protein levels of androgen receptor (AR), p-Akt and PAX7. Furthermore, T treatment significantly promoted myoblasts cultured in vitro entering a new cell cycle and increased PAX7-positive cells. The mRNA and protein expression of AR and PAX7 were upregulated when treated with T compared to that of the control. The addition of T induced proliferation accompanied by increasing AR level as well as PI3K (Phosphoinositide 3-kinase)/Akt activation. However, T-induced proliferation was attenuated by AR, PI3K, and Akt-specific inhibitors. These data indicated that the pro-proliferative effect of T was regulated though AR in response to the activation of PI3K/Akt signalling pathway.

Bisphenol A (BPA) as an endocrine-disrupting chemical with weak estrogenic activity affects formation of primordial follicles. This study aimed to identify the potential effects and molecular mechanisms of BPA on meiosis and primordial follicle formation in chickens. The results suggest that the cortical layer was thickened and the number of germ cells that entered into meiosis was increased in BPA-treated ovaries. The percentage of γH2AX-positive cells increased significantly. In addition, up-regulated mRNA expression of meiotic genes, including stimulated by retinoic acid gene 8 (Stra8), disrupted meiotic cDNA 1 homologue (Dmc1) and synaptonemal complex protein 3 (Scp3) were observed in BPA-treated ovaries. Therefore, progression to meiosis prophase I was accelerated by exposure to BPA. Furthermore, the results demonstrated that injection of BPA resulted in hypomethylation of Dazl (Deleted in A Zoospermia-Like gene) and Stra8 and up-regulation mRNA expression of Dazl and Stra8 during meiotic onset. Finally, the relationship between estrogen receptor (ER) expression and BPA-induced meiosis was revealed using an in vitro ovarian culture system. BPA enhanced ERβ expression at the levels of mRNA and protein, while BPA exerted no significant effect on ERα and membrane-bound estrogen receptor (GPR30) expression. The inducing effects of BPA on meiosis were blocked by ER inhibitor. Collectively, these results demonstrate the dynamic ovarian response to BPA exposure, which indicate that BPA affects the formation of primordial follicles by promoting meiotic progression of oocytes via hypomethylation of Dazl and Stra8 and ERβ signaling pathways.

Background Circular RNAs (circRNAs) belong to a new type of endogenous non-coding RNA and plays a key role in carcinogenesis. Circ-ZKSCAN1 (hsa_circ_0001727) has been proven to be a tumor-dependent circRNA. However, its role in non-small cell lung cancer (NSCLC) has been underreported. Methods The expression patterns of circ-ZKSCAN1 were determined using qRT-PCR in NSCLC samples and cell lines. Cell proliferation was examined utilizing the CCK-8 assay. Cell migration and invasion were evaluated using the Transwell assay. The combination of circ-ZKSCAN1 and miR-330-5p in NSCLC cells was analyzed by RNA pull-down and luciferase reporter assay. We used the bioinformatics software circbank, CircInteractome, TargetScan and Miranda to predict circRNA-miRNA and miRNA-mRNA interactions. Results Our results showed that circ-ZKSCAN1 was significantly up-regulated in NSCLC, closely related to malignant characteristics and poor prognosis, and clinically related to tumor size and clinical stage. Subsequent experiments showed that circ-ZKSCAN1 could inhibit the growth of NSCLC cells in vitro and in vivo. Importantly, circ-ZKSCAN1 can act as a sponge of carcinogenic miR-330-5p to increase the expression of FAM83A, resulting in the inhibition of MAPK signal transduction pathway, thus promoting the progress of NSCLC. Interestingly, the increase in FAM83A expression caused by circ-ZKSCAN1 overexpression could in turn promote the expression of circ-ZKSCAN1. Conclusions Circ-ZKSCAN1 is a key positive regulator of NSCLC, and clarifies the potential molecular mechanism of the new circ-ZKSCAN1/miR-330-5p/FAM83A feedback loop in promoting the progress of NSCLC.

Skeletal muscle development is an orchestrated progress that is primarily regulated by temporospatial expression of myogenic regulatory factors (MRFs). Recent studies demonstrated that DNA demethylation also exerted a critical role in myogenesis. However, the function of Tet2 in the regulation of chicken myogenesis still remains unknown. In the present study, the role of Tet2 in regulating myogenic differentiation was determined by using a model of primary myoblasts from chickens. The expression of Tet2 was significantly elevated during myoblast differentiation. Meanwhile, the level of 5hmC in genomic DNA was increased, but H3K9me2 and H3K27me3 were markedly reduced following differentiation. Knockdown of Tet2 significantly inhibited the formation of multinucleated myotubes, which was accompanied by a reduction of relevant pivotal MRFs. Moreover, the level of 5hmC decreased sharply in Tet2 knockdown myoblasts. Attenuated differentiated myoblasts that resulted from reduced Tet2 also demonstrated an increased level of H3K9me2 and H3K27me3. Collectively, these results indicated that Tet2 played an essential role during myogenesis, which affected demethylation of genomic DNA and histone to regulate expression of MRFs and therefore, contributed to myoblast differentiation.

Additional file 2: Figure S1. The methylmix model of the rest of methylation-driven genes in ESCC.

High-quality chicken meat is an important source of animal protein for humans. Gene expression profiles in breast muscle tissue were determined, aiming to explore the common regulatory genes relevant to muscle and intramuscular fat (IMF) during the developmental stage in chickens. Results show that breast muscle weight (BMW), breast meat percentage (BMP, %), and IMF (%) continuously increased with development. A total of 256 common differentially expressed genes (DEGs) during the developmental stage were screened. Among them, some genes related to muscle fiber hypertrophy were upregulated (e.g., CSRP3, LMOD2, MUSTN1, MYBPC1), but others (e.g., ACTC1, MYL1, MYL4) were downregulated from Week 3 to Week 18. During this period, expression of some DEGs related to the cells cycle (e.g., CCNB3, CCNE2, CDC20, MCM2) changed in a way that genetically suggests possible inhibitory regulation on cells number. In addition, DEGs associated with energy metabolism (e.g., ACOT9, CETP, LPIN1, DGAT2, RBP7, FBP1, PHKA1) were found to regulate IMF deposition. Our data identified and provide new insights into the common regulatory genes related to muscle growth, cell proliferation, and energy metabolism at the developmental stage in chickens.

Ten–eleven translocation 1 (Tet1) is involved in DNA demethylation in primordial germ cells (PGCs); however, the precise regulatory mechanism remains unclear. In the present study the dynamics of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) in developing PGCs and the role of Tet1 in PGC demethylation were analysed. Results show that 5mC levels dropped significantly after embryonic Day 4 (E4) and 5hmC levels increased reaching a peak at E5–E5.5. Interestingly, TET1 protein was highly expressed during E5 to E5.5, which showed a consistent trend with 5hmC. The expression of pluripotency-associated genes (Nanog, PouV and SRY-box 2 (Sox2)) and germ cell-specific genes (caveolin 1 (Cav1), piwi-like RNA-mediated gene silencing 1 (Piwi1) and deleted in azoospermia-like (Dazl)) was upregulated after E5, whereas the expression of genes from the DNA methyltransferase family was decreased. Moreover, the Dazl gene was highly methylated in early PGCs and then gradually hypomethylated. Knockdown of Tet1 showed impaired survival and proliferation of PGCs, as well as increased 5mC levels and reduced 5hmC levels. Further analysis showed that knockdown of Tet1 led to elevated DNA methylation levels of Dazl and downregulated gene expression including Dazl. Thus, this study reveals the dynamic epigenetic reprogramming of chicken PGCs invivo and the molecular mechanism of Tet1 in regulating genomic DNA demethylation and hypomethylation of Dazl during PGC development.

During embryonic development, IGF-1 fulfils crucial roles in skeletal myogenesis. However, the involvement of IGF-1-induced myoblast proliferation in muscle growth is still unclear. In the present study, we have characterised the role of IGF-1 in myoblast proliferation both in vitro and in vivo and have revealed novel details of how exogenous IGF-1 influences myogenic genes in chicken embryos. The results show that IGF-1 significantly induces the proliferation of cultured myoblasts in a dose-dependent manner. Additionally, the IGF-1 treatment significantly promoted myoblasts entering a new cell cycle and increasing the mRNA expression levels of cell cycle-dependent genes. However, these effects were inhibited by the PI3K inhibitor LY294002 and the Akt inhibitor KP372-1. These data indicated that the pro-proliferative effect of IGF-1 was mediated in response to the PI3K/Akt signalling pathway. Moreover, we also showed that exogenous IGF-1 stimulated myoblast proliferation in vivo. IGF-1 administration obviously promoted the incorporation of BrdU and remarkably increased the number of PAX7-positive cells in the skeletal muscle of chicken embryos. Administration of IGF-1 also significantly induced the upregulation of myogenic factors gene, the enhancement of c-Myc and the inhibition of myostatin (Mstn) expression. These findings demonstrate that IGF-1 has strong activity as a promoter of myoblast expansion and muscle fiber formation during early myogenesis. Therefore, this study offers insight into the mechanisms responsible for IGF-1-mediated stimulation of embryonic skeletal muscle development, which could have important implications for the improvement of chicken meat production.

Many genes participate in the process of ovarian germ cell development, while the combined action mechanisms of these molecular regulators still need clarification. The present study was focused on determination of differentially expressed genes and gene functions at four critical time points in chicken ovarian development. Comparative transcriptional profiling of ovaries from embryonic day 5.5 (E5.5), E12.5, E15.5 and E18.5 was performed using an Affymetrix GeneChip chicken genome microarray. Differential expression patterns for genes specifically depleted and enriched in each stage were identified. The results showed that most of the up- and downregulated genes were involved in the metabolism of retinoic acid (RA) and synthesis of hormones. Among them, a higher number of up- and downregulated genes in the E15.5 ovary were identified as being involved in steroid biosynthesis and retinol metabolism, respectively. To validate gene changes, expressions of twelve candidate genes related to germ cell development were examined by real-time PCR and found to be consistent with the of GeneChip data. Moreover, the immunostaining results suggested that ovarian development during different stages was regulated by different genes. Furthermore, a Raldh2 knockdown chicken model was produced to investigate the fundamental role of Raldh2 in meiosis initiation. It was found that meiosis occurred abnormally in Raldh2 knockdown ovaries, but the inhibitory effect on meiosis was reversed by the addition of exogenous RA. This study offers insights into the profile of gene expression and mechanisms regulating ovarian development, especially the notable role of Raldh2 in meiosis initiation in the chicken.

During embryonic development, IGF-1 fulfils crucial roles in skeletal myogenesis. However, the involvement of IGF-1-induced myoblast proliferation in muscle growth is still unclear. In the present study, we have characterised the role of IGF-1 in myoblast proliferation both in vitro and in vivo and have revealed novel details of how exogenous IGF-1 influences myogenic genes in chicken embryos. The results show that IGF-1 significantly induces the proliferation of cultured myoblasts in a dose-dependent manner. Additionally, the IGF-1 treatment significantly promoted myoblasts entering a new cell cycle and increasing the mRNA expression levels of cell cycle-dependent genes. However, these effects were inhibited by the PI3K inhibitor LY294002 and the Akt inhibitor KP372-1. These data indicated that the pro-proliferative effect of IGF-1 was mediated in response to the PI3K/Akt signalling pathway. Moreover, we also showed that exogenous IGF-1 stimulated myoblast proliferation in vivo. IGF-1 administration obviously promoted the incorporation of BrdU and remarkably increased the number of PAX7-positive cells in the skeletal muscle of chicken embryos. Administration of IGF-1 also significantly induced the upregulation of myogenic factors gene, the enhancement of c-Myc and the inhibition of myostatin (Mstn) expression. These findings demonstrate that IGF-1 has strong activity as a promoter of myoblast expansion and muscle fiber formation during early myogenesis. Therefore, this study offers insight into the mechanisms responsible for IGF-1-mediated stimulation of embryonic skeletal muscle development, which could have important implications for the improvement of chicken meat production.