Abstract:Objective：To investigate whether alternative splicing regulate the stage transition of follicle assembly in mice，the transcriptome data of single germ cells in new born mouse ovary were analyzed. Methods：Bioinformatics analysis were performed on transcriptome data of single germ cells in mice to detect alternative splicing events in three cell development stages during follicle assembly and differential alternative splicing events between stage transitions. PCR was used to validate dynamic stage transition related isoform expression. Results：The current study found abundant differential alternative splicing events between stage transitions during follicle assembly in mice. The most enriched splicing events were detected in follicle stage，also the most enriched differential splicing events were detected between cyst break down and follicle stage. The results of PCR validated the presence of isoforms of meiosis related genes and showed their shifts of alternative isoform usage at stage transitions. Conclusion：Alternative splicing was associated with cell development stage transitions during follicle formation in mice，regulating the transformation from germ cell to oocytes.

Abstract:Objective：To investigate the differences of cardiovascular development in zebrafish after knockout of the hemoglobin gene on chromosome 3 and the underlying mechanism. Methods：We used CRISPR/Cas9 technology to knock out the fragment from hbba2 to hbae3 of the zebrafish hemoglobin gene on chromosome 3 to obtain heterozygous zebrafish（Hb ^+/-）. The hemoglobin content and distribution of Hb ^+/- and wild-type zebrafish（WT）fertilized eggs were detected，the development of blood vessels was observed， and the number of 5-month-old erythrocytes of both were counted. The transcriptome of the heart of adult zebrafishes were analyzed by sequencing and validated by qRT-PCR. Results：A significantly reduced hemoglobin content in Hb ^+/- embryos and ectopic in 50 hpf embryos was detected. Compared with WT，Hb^+/- had a retarded development of abdominal major vessels and common cardinal veins in the abdomen，a more rapid development of the subintestinal veins，and hyperplasia of the caudal intersegmental vessels. There was no significant difference in erythrocytes between Hb ^+/- and adult WT（P > 0.05）. Transcriptome analysis of the heart of adult zebrafish revealed that the decreased hemoglobin levels led to the up-regulation of nos2b and cybb，down-regulation of hif3a，and activation of compensatory protective mechanisms in mitochondria. Conclusion：Hemoglobin gene knockout affects hemoglobin levels and vascular development in embryonic zebrafish，and the normal physiological activity of adult fish can be maintained by regulating gene expression and mitochondrial compensation mechanism.

Abstract:Objective：To investigate the relationship between acidic deoxycholic acid induced mitochondrial DNA（mtDNA） damage and release of human esophageal epithelial cells and cGAS -STING pathway in the development of esophageal epithelial cell inflammation. Methods：HEECs were divided into control group and acidic deoxycholic acid treatment group. The viability of cells was measured by CCK -8 assay. Changes of reactive oxygen species，mitochondrial reactive oxygen species and mitochondrial membrane potential were detected by fluorescence microscope and flow cytometry. ATP was detected by the luminometer. The ultrastructure of mitochondria was observed by transmission electron microscope. The mtDNA copy number was evaluated by qPCR. The expressions of γH2AX，cGAS，STING，p -NF -κB p65 and NF -κB p65 were detected by Western blotting. The mRNA expressions of inflammatory cytokines IL-6 and IL-1β were detected by qPCR. Results：CCK-8 assay showed that the viability of cells treated with acidic deoxycholic acid decreased in a dose-time dependent manner. The production of intracellular ROS and mtROS increased，while MMP and ATP decreased. Compared with the control group，the expression of γH2AX increased after acidic deoxycholic acid，mtDNA released into the cytoplasm，mtDNA copy number reduced，the expressions of cGAS，STING and p-NF-κB p65 were increased，and the expressions of inflammatory cytokines IL-6 and IL-1β were elevated. After pretreatment with cGAS inhibitor RU.521，the expression levels of cGAS and STING were inhibited and the expression of p-NF-κB p65 was partially inhibited，and the levels of inflammatory cytokines IL-6 and IL-1β were decreased. Conclusion：The in vitro experiments have shown that acidic deoxycholic acid can induce mitochondrial dysfunction，mitochondrial DNA damage and release，and mediate HEEC inflammation. The mechanism may be related to the activation of cGAS-STING pathway.

Abstract:Objective：Based on the glycosylation related gene set，the TCGA-GTEx joint public database was used to identify genes significantly related to the occurrence and poor prognosis of pancreatic cancer，and the tissue samples of our center and in vitro experiments were used for verification. Methods：The glycosylation gene B4GALT5 related to the risk and poor prognosis of pancreatic cancer was screened through the public database，and was verified using the tissue samples of our center with clinical and prognostic information. We constructed pancreatic cancer cell lines with B4GALT5 interference and overexpression，and observed the effects of B4GALT5 on the malignant biological behavior of pancreatic cancer cells in vitro through CCK -8，wound healing，transwell invasion experiments. Results：The expression of B4GALT5 in pancreatic cancer tissue was significantly higher than that in normal pancreatic tissue. The increased expression of B4GALT5 was positively correlated with the risk and poor prognosis of pancreatic cancer，and negatively correlated with the infiltration of CD4 ⁺ T cells and NK T cells. B4GALT5 played a role in exacerbating the proliferation， movement and invasion of pancreatic cancer cells in vitro. Conclusion：The glycosylation gene B4GALT5 can promote the malignant progression of pancreatic cancer，and may be used as a new biomarker to indicate the risk and poor prognosis of pancreatic cancer.

Abstract:Objective：To explore the effects of meningeal lymphatic vessel transport dysfunction on lipopolysaccharide（LPS）-induced central nervous system inflammation in mice. Methods：Firstly，sixteen C57BL/6 mice were randomly divided into four groups， normal saline was injected intraperitoneally in the Control group. The LPS groups were injected intraperitoneally with LPS（2 mg/kg）， and then the transport function of meningeal lymphatic vessels，activation of microglia and levels of interleukin-6（IL-6）and interleukin-1β （IL -1β）in the hippocampus were observed after 12 h，24 h and 72 h. Secondly，eight mice were randomly assigned to two groups： Control group and VEGFR3 inhibitor MAZ51 group，to observe the effects of MAZ51 on the transport function of meningeal lymphatic vessels. Finally，twenty-four mice were divided into four groups as follows：control group，MAZ51 group，LPS group，and LPS+MAZ51 group. MAZ51（10 mg/kg）was preinjected intraperitoneally into MAZ51 group and LPS+MAZ51 group for five days per week with a total of 30 days. After six weeks，LPS was injected into LPS group and LPS+MAZ51 group. A day later，behavioral experiments that assess the ability of mice to escape from fear were conducted；the activation of microglia in the hippocampus was measured by immunohistochemistry；the expression of IL-6 and IL-1β was evaluated by ELISA method. Results：The area of LYVE-1 in meninges and the area of OVA-647 in the deep cervical lymph nodes were significantly decreased 24 h after intraperitoneal injection of LPS（P < 0.01），and were lower than the basal level for 72 h（P < 0.01）. The activation of microglia and levels of IL-6 and IL-1β in the hippocampus were significantly increased after 24 h（P < 0.01）. Compared with the control group，the area of OVA-647 fluorescence in the MAZ51 group was significantly reduced（P < 0.01）. The activation of microglia and the expression of inflammatory factors in the hippocampus of mice in the LPS+MAZ51 group were increased（P < 0.01）and the freezing time was significantly reduced（P < 0.01）. Conclusion：The impairing transport function of meningeal lymphatic vessels aggravates LPS-induced central inflammation and cognitive dysfunction in mice by increasing inflammatory mediator accumulation and activating microglia.

Abstract:Objective：To investigate the association between vitamin D（VD）deficiency and the pathogenesis of polycystic ovarian syndrome（PCOS）. Methods：Vitamin D levels were detected in 34 patients with PCOS and 30 healthy women，and the correlation analysis was performed. Vitamin D deficiency mouse model and PCOS mouse model were constructed and divided into four groups： control group（CTR group），PCOS group，vitamin D deficiency group（VD^- group），and vitamin D deficiency combined with PCOS group （VD^- + PCOS group）. Estrous cycle and ovary morphological changes were observed. Serum sex hormone indexes and glucose lipid metabolism indexes were detected. Real-time quantitative PCR（RT-qPCR）was used to detect the mRNA levels of hormone synthase and sex hormone receptor in ovarian tissues. Results：The levels of serum 25- hydroxyvitamin D［25（OH）D］in PCOS women were significantly lower than those in normal healthy women［（16.49 ± 6.50）ng/mL vs.（20.08 ± 5.28）ng/mL，P=0.019］. The levels of luteinizing hormone（LH），testosterone（TT），LH/FSH ratio and free androgen index（FAI）in PCOS group were significantly higher than those in control group，while the levels of sex hormone binding globulin（SHBG）were significantly lower than those in control group，and the difference was statistically significant（P < 0.05）. Correlation analysis showed that serum 25（OH）D levels of the total sample population were negatively correlated with LH（r=-0.271，P < 0.05），LH/FSH ratio（r=-0.314，P < 0.05），TT（r=-0.276，P < 0.05），prolactin（PRL）（r=-0.274，P < 0.05）and FAI（r=-0.312，P < 0.05）respectively，and serum 25（OH）D levels in PCOS patients were negatively correlated with PRL（r=-0.404，P < 0.05）. Vitamin D deficiency stalled the estrous cycle in mice. Compared with the CTR group，the serum LH levels of mice in other three groups were significantly increased（P < 0.05）. Compared with the CTR group， the mRNA level of androgen receptor（AR）in ovarian tissues of mice in VD^- group was increased（P < 0.05）. Conclusion：Vitamin D deficiency existed in patients with PCOS. There was a negative correlation between serum 25（OH）D and PRL in PCOS patients. Vitamin D deficiency disturbs normal estrous cycle and affects sex hormone levels in mice. Vitamin D deficiency may participate in the occurrence of PCOS by regulating the expression of sex hormone receptor.

Abstract:Objective：The current study aims to investigate the relationship between hsa_circ_0005389 and neonatal acute respiratory distress syndrome（NARDS）based on high-throughput sequencing and bioinformatics analysis of human circular RNA （circRNA）to provide a new direction for the diagnosis and treatment of NARDS. Methods：The acute lung injury model was induced by lipopolysaccharide（LPS）. The expression of inflammatory markers and related pathway indexes were detected by RT-qPCR and Western blot in the negative control group and the intervention group with knockdown of hsa_circ_0005389 expression. CCK -8 and flow cytometry were used to detect the proliferation and apoptosis of blank A549 cells and A549 cells with establishment of acute lung injury model，both of which were knocked-down or over-expressed hsa_circ_0005389. Result：The mRNA relative expression and the protein expression of inflammatory markers and related pathway indexes were highly expressed and increased significantly after exposed to 10 μg/cm² LPS for 48 h（P < 0.05）. Compared with the negative control group，the relative mRNA expression level of soluble tumor necrosis factor receptor 1（sTNFR1）and the protein expression of tumor necrosis factor α（TNF-α）in A549 cells were significantly decreased（P < 0.05）after knocking down the expression of hsa_circ_0005389. Inflammatory markers and related pathway indexes decreased significantly（P < 0.001）in the acute lung injury model after knocking down the express of hsa_circ_ 0005389. The proliferation of A549 cells were accelerated and the apoptosis rate was decreased（P < 0.05）when the expression of hsa_circ_0005389 was down-regulated，while the proliferation of A549 cells was slowed down and the apoptosis rate was increased after hsa_circ_0005389 expression was over-regulated（P < 0.05）by using CCK -8 and flow cytometry as compared with the negative control group. Conclusion：In acute lung injury cell model，hsa_circ_0005389 promoted the expression of inflammatory markers of lung injury，affecting the proliferation and apoptosis of lung epithelial cells. These results suggested that hsa_circ_0005389 was involved in the inflammatory process of NARDS，which may be a potential intervention target.

Abstract:Objective：To establish a new pancreatic cancer risk assessment model based on the differential expression of m1A-related genes in multiple databases and clinical parameters，and provide theoretical basis for pancreatic cancer treatment and prognosis analysis. Methods：Differential analysis of m1A-related genes was performed in the TCGA -GTEx database. Cox analysis and LASSO regression were used to construct a pancreatic cancer prognosis risk model，and the accuracy and sensitivity of the model were verified by gene expression and clinical data from the GEO database and pancreatic cancer cases in our center. Results：CRLS1 and C7orf50 were selected to form the risk model. The study systematically verified that the model had significant efficacy in indicating the prognosis of pancreatic cancer，and the model combined with tumor mutation burden（TMB）had a stronger prognostic indication ability. Conclusion：A new risk model is constructed to indicate the prognosis of pancreatic cancer patients.

Abstract:Objective：To screen genes related to oxaliplatin resistance in colon cancer and verify the influence of them on the oxaliplatin treatment of colon cancer. Methods：Data set GSE42387 was downloaded from NCBI-GEO and analyzed comprehensively， and oxaliplatin resistance genes were screened. Ualcan and GEPIA databases were used to detect the expression of drug-resistant genes in colon cancer and their influence on prognosis. Transfection of siRNA-ZIP8 interfered with zinc transporter 8（ZIP8）expression in colon cancer cell SW620. CCK-8 proliferation assay detected the effect of oxaliplatin on colon cancer cell proliferation inhibition after ZIP8 expression was inhibited. Flow cytometry was used to detect the effect of oxaliplatin on colon cancer cell apoptosis after the expression of ZIP8 was decreased. Immunohistochemical staining was performed on colon cancer tissue samples before chemotherapy to evaluate the relationship between ZIP8 and chemotherapy effect. Results：Four drug-resistant genes（ZIP8，NUPR1，SLC43A1 and TEME73）were selected. ZIP8 was highly expressed in colon cancer tissues（P < 0.05）and correlated with patient prognosis（P= 0.005）. Under the same drug concentration，the proliferation inhibition rate of the siZIP8 group was significantly lower than that in the siNC group，the half inhibitory concentration of oxaliplatin in the siZIP8 group was lower than that in the siNC group（P < 0.05）. Patients with low ZIP8 expression had longer overall survival（P=0.001）and disease -free survival（P < 0.001）. Conclusion：ZIP8 promotes oxaliplatin resistance in colon cancer patients and can be used as an indicator for the prognosis of chemotherapy.

Abstract:Objective：To investigate the impact of the graft content including mononuclear cells（MNC），CD34⁺ cells and T-lymphocyte subsets on post-transplant survival. Methods：A retrospective analysis of 121 patients with hematologic malignant diseases undergoing allogeneic hematopoietic stem cell transplantation（allo -HSCT）was performed to investigate the effects of the doses of MNC，CD34 ⁺ cells，CD3⁺ T cells，CD4⁺ T cells，CD8⁺ T cells and regulatory T（Treg）cells in the grafts on hematopoietic reconstitution and survival. Results：Neutrophils were successfully engrafted in 120 patients，and patients with high doses of CD34⁺ cells（≥6.90×10⁶ /kg）（P < 0.001）， CD3⁺ T cells（≥6.24×10⁸ /kg）（P=0.042）and CD8⁺ T cells（≥1.05×10⁸ /kg）（P=0.021）had more rapid engraftment of neutrophils. Platelets were successfully engrafted in 119 cases，and high dose CD34 ⁺ cells were associated with faster platelet reconstruction（P=0.001）. Acute graft-versus-host disease（aGVHD）occurred in 53（43.8%）patients and 28 patients had chronic graft-versus-host disease （cGVHD）. One and 3-year overall survival（OS）rates were 83.5% and 68.0%，and one and 3-year progression -free survival（PFS） rates were 75.0% and 64.4%. In univariate analysis，the patients with high MNC dose（≥9.79×10⁸ /kg），high CD3 ⁺ T cells dose and CD4⁺ /CD8⁺ T cells <3.57 had better OS and the high MNC group had better PFS（P=0.061）. Multivariate analysis showed that the CD4⁺ / CD8⁺ T-cell < 3.57 group had better OS（HR=0.288，95%CI：0.084~0.988，P=0.048）. One and 3-year cumulative incidence of relapse（CIR）were 18.2% and 26.8%，respectively. Patients in the higher MNC and CD8 ⁺ T -cell groups had lower CIR. Conclusion：Graft components has an important impact on the prognosis of allo-HSCT. High doses of CD34⁺ cells promote faster platelet implantation, and patients with a CD4⁺ /CD8⁺ T-cell ratio < 3.57 in the graft have better OS and lower CIR.

Abstract:Objective：This article aims to use computer model tools to discuss transcranial direct current stimulation（tDCS）for clinical treatment problems，applying ROAST（realistic volumetric-approach to simulate transcranial electric stimulation）to the analysis of the influence of electrode position errors in the tDCS clinical treatment，and quantitatively analyze effects from the position offsets. Methods：This study takes F3 and F4 as an example to treat depression，simulates and calculates the electric field dose excited in the intracranial target area，compares the influence of the electrode position errors on the intracranial treatment target electric field dose，and gives the electric field dose variation curve under application of 2 mA and 4 mA current. Results：The simulation shows among 32 kinds of physical position errors，the pad electrode and the disk electrode cause the electric field dose to vary from-10.3% to 72.4%，-12.9% to 11.3%，both decreasing and increasing；this result is due to the non -uniformity of the cerebral cortex. When the electric field dose is reduced，the electric field in intracranial target area will be low than the effective electric field dose，so it is possible that the clinical treatment effect of tDCS may not be good. The simulation also shows that when 4.0 mA is applied，the intracranial target area will produce an effective electric field dose. In addition，the variation of the electric field dose generated by 32 kinds of error positions in the intracranial target area is basically the same as that when 2 mA is applied. The degree of electric field dose change is consistent with the magnitude of the applied current It’s roughly okay. The degree of electric field dose change is roughly irrelevant to the magnitude of the applied current. Conclusion：ROAST can be used to find the individual dose optimization of tDCS for clinical doctors.

Abstract:Polycyclic aromatic hydrocarbons（PAHs）are a large group of persistent organic pollutants that widely exist in the environment. The risk of PAHs exposure has increased with the increasing industrialization and fossil fuels consumption in China. It has been reported that PAHs are potential risk factors for asthma in children，but the specific pathogenesis has not been fully clarified. In recent years，the development of metabolomics has provided a promising approach to elucidate the toxicological mechanism of environmental pollutants. In this paper，we will review the mechanism of PAHs mediating inflammation and inducing childhood asthma by changing epigenetic patterns and activating aromatic hydrocarbon receptor signaling pathway from the perspective of metabolism， especially one carbon metabolism and tryptophan metabolism.

Abstract:Chronic obstructive pulmonary diseases（COPD）is a kind of disease characterized by airflow restriction. It is related to complex pathological changes caused by inhalation of harmful gases or particles such as cigarettes and smoke，including inflammation of large and small airways and destruction of lung parenchyma. The pathogenesis of COPD is still unclear，and is currently believed to be related to airway inflammation，immune factors，oxidative stress，protease and antiprotease imbalance. Molecular biomarkers closely related to the pathogenesis of COPD play an important role in the occurrence and development of the disease. This paper reviewed the molecular biomarkers corresponding to different pathogenesis of COPD.

Abstract:Complex regional pain syndrome is one of the neuropathic pain disorders，and its pathogenesis is still unclear. At present， it is believed that the etiology of CRPS is multifactorial. Early diagnosis and treatment can improve its prognosis. This article reviews the latest research progress in the epidemiology，pathogenesis，clinical diagnosis，and treatment of CRPS.

Abstract:With changes in life style and dietary structure，the global prevalence of nonalcoholic fatty liver disease（NAFLD）is increasing year by year，posing a serious burden on health care systems. While clinicians’understanding of the pathophysiological mechanisms of NAFLD continues to improve，effective treatments are still being explored，and life style interventions remain the first- line therapy for NAFLD as recommended by numerous guidelines and physicians. Sleep is a complex，highly regulated process that is critical to human health. There is a growing consensus that sleep disorders are closely associated with chronic liver disease，especially NAFLD. However，it is not clear how sleep disorders affect NAFLD. Therefore，the purpose of this paper is to review the relationship between sleep disorders and NAFLD and to enable NAFLD patients to make life style changes，which will benefit NAFLD patients.

Abstract:Ischemic stroke is considered to be one of the major causes of death and disability worldwidely，and is correlated to a series of complex pathophysiological processes. However，the time window of thrombolysis and thrombectomy commonly used in clinic is relatively short，and the treatment still has limitations. Oleanolic acid is a triterpenoid compound that has antioxidative，anti-neuroinflammatory and other functions. It exists widely in various precious traditional Chinese medicines. Existing studies have proved that oleanolic acid can reduce excitotoxicity，Ca²⁺ overload，mitochondrial dysfunction and blood -brain barrier permeability，and can anti -inflammation，anti oxidation，anti-apoptosis and improve angiogenesis and neurogenesis. This manuscript reviews the role and mechanism of oleanolic acid in the treatment of ischemic stroke，providing a reference for its clinical application.