[Bioinformatics analysis of primary biliary cholangitis key genes and molecular mechanisms].

Objective: To extract the differentially expressed key genes of primary biliary cholangitis (PBC) using bioinformatics methods, so as to provide information for further study into the mechanism. Methods: The GSE119600 dataset was downloaded from the GEO database to obtain differentially expressed genes. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed for differentially expressed genes. Protein-protein interaction (PPI) network reconstruction, Cytoscape software visualization, and core gene screening were performed. The area under the receiver operating characteristic curve (ROC AUC) was used to assess the diagnostic effectiveness of genes and plot the pROC software package. The x-Cell software was used to calculate the enrichment score of 34 immune cells in each sample. Finally, four key genes (PSMA4, PSMA1, PSMB1, and PSMA3) were selected. Blood samples were analyzed using the qPCR method. Results:: A total of 373 immune-related differentially expressed genes were identified. Eight genes (PSMC6, PSMB2, PSMB1, PSMA3, PSMA4, PSMA1, PSMD7, and PSMB5) were screened from the 178 nodes and 596 edges as hub genes of the PPI network, which were significantly related to amino acid metabolism, hematopoietic stem cell differentiation, cell cycle, and immune processes. PSMA4, PSMA1, PSMB1, and PSMA3 were defined as immunological biomarkers for PBC with an AUC value of the ROC curve > 0.7. Immunoinfiltrating cell analysis showed that the proportion of eosinophils was significantly higher in PBC patients compared to the control group, whereas the proportion of CD4+ memory T cells, plasma cells, Th2 cells, and cDC cells was significantly lower in PBC patients than the control group. Plasma cells were associated with all four immunological biomarkers. Seven PBC patients and seven healthy subjects were selected for peripheral blood qPCR validation, which demonstrates that PSMB1, PSMA3, PSMA1, and PSMA4 levels were significantly lower in PBC patients than healthy subjects, with a statistically significant difference. Conclusion:: Bioinformatics screened eight key genes, of which four were key immunological markers and may serve as a basis for clinical diagnosis and mechanism exploration.

[Clinical study of digital six-axis external fixation frame based on CT data for tibiofibular fractures].

Objective: To investigate the clinical effect of applying the digital six-axis external fixation frame based on CT data in the treatment of tibiofibular fractures. Methods: The clinical data of 43 patients with tibiofibular fractures treated by the self-developed digital six-axis external fixation frame based on CT data at Integrated Orthopedic Department of Traditional Chinese Medicine (TCM) and Western Medicine,HongHui Hospital from January 2018 to January 2021 were retrospective analysis.There were 27 males and 16 females,aged (36.0±9.4) years(range:25 to 50 years).AO classification:15 cases of 42A,11 cases of 42B, and 17 cases of 42C.There were 7 open fractures and Gustilo fracture classification:2 cases of type Ⅰ,4 cases of type Ⅱ,and 1 case of type Ⅲ.The two or three plane rings were connected with six connecting rods to form a complete six-axis external fixation frame,and the distal and proximal fracture blocks were connected to the distal and proximal rings by fixation pins,and the lengths of the six connecting rods needed to be adjusted were calculated by using the supporting software according to the CT data after surgery,and then the lengths of the connecting rods were adjusted one by one to complete the reduction of the fracture. The reduction accuracy of this six-axis external fixation brace was evaluated by measuring postoperative radiographs; postoperative recovery and complications were collected,the time of brace removal was recorded,and the function of the affected limb was evaluated according to the Johner-Wruhs score at the final follow-up. Results: Postoperative radiographs showed that all patients achieved satisfactory reduction with lateral displacement(M(IQR)) of 2.3(2.5) mm (range:0.3 to 7.3 mm),anteroposterior displacement of 2.1 (2.4) mm (range:0.3 to 5.7 mm),anteroposterior angulation of 2.5(2.4)°(range:0 to 5°),internal and external angulation of 2.1(1.5)°(range:0 to 4°), and no significant internal or external rotational deformity was detected on the exterior.On the second postoperative day,all patients were able to walk with partial weight-bearing on crutches. All 43 patients were followed up for more than 6 months,with a follow-up period of (33.3±7.3) weeks (range:24 to 42 weeks).The external fixation frame was removed after the fracture healed.The external frame was removed at 20(3)weeks (range:18 to 25 weeks) postoperatively. Up to the final follow up, no secondary fracture occurred in any of them.The Johner-Wruhs score of the affected limb at the last follow-up was excellent in 39 cases and good in 4 cases. Conclusion: The digital six-axis external fixator based on CT data for tibiofibular fractures has the advantages of precise reduction,firm fixation,simple operation,rapid fracture healing,and minimal trauma, which is a minimally invasive method for treating tibiofibular fractures,especially suitable for patients with poor skin and soft tissue conditions such as open injuries.

[Safety and efficacy of transcatheter tricuspid valve replacement with LuX-Valve in patients with severe tricuspid regurgitation].

Objectives: To evaluate the safety and efficacy of LuX-Valve on the treatment of severe tricuspid regurgitation (TR). Methods: This is a prospective observational study. From September 2018 to March 2019, 12 patients with severe TR, who were not suitable for surgery, received LuX-Valve implantation in Changhai Hospital. LuX-Valve was implanted under general anesthesia and the guidance of transesophageal echocardiography and X-ray fluoroscopy. Access to the tricuspid valve was achieved via a minimally invasive thoracotomy and transatrial approach. Main endpoints were surgery success and device success. Surgery success was defined as successful implanting the device and withdrawing the delivery system, positioning the valve correctly and stably without severe or life-threatening adverse events. Device success was defined as satisfied valve function (TR severity reduction ≥ 2 grades, tricuspid gradient ≤ 6 mmHg (1 mmHg=0.133 kPa)), absence of malposition, valve failure and reintervention, major adverse events including device related mortality, embolization, conduction system disturbances and new onset shunt across ventricular septum at day 30 post implantation. Results: A total of 12 patients with severe to torrential TR were included in this study. The age was (68.5±6.9) years and 7 were female. All patients had typical right heart failure symptoms. Procedural success was achieved in all cases, there was no intraprocedural mortality or transfer to open surgery. TR significantly improved after LuX-Valve implantation (none/trivial in 8 patients, mild in 3 patients and moderate in 1 patient). The average device time was (9.2±4.2) minutes. Intensive care unit duration was 3.0 (2.0, 4.8) days. One patient died at postoperative day 18 due to non-surgery and device reasons. Transthoracic echocardiography at 30 days after operation showed that TR was significantly reduced (none/trivial in 8 patients, mild in 2 patients and moderate in 1 patient) and device success was achieved in 11 cases. All survived patients experienced a significant improvement in life quality with significantly improvement in New York Heart Association (NYHA) classification (Ⅰ and Ⅱ: 6/11 post operation vs. 0/11 before operation, P=0.012) and there were no device related complications in this patient cohort. Conclusions: LuX-Valve implantation is feasible, safe and effective for the treatment of patients with severe TR.

[Application of a new type of angle-adjustable osteotomy guide in closing wedge distal femoral osteotomy].

Objective: To explore the feasibility of fast and accurate osteotomy using a new angle adjustable osteotomy guide (AAOG) in closing wedge distal femoral osteotomy(CWDFO). Methods: The clinical data of 14 patients (17 knees) with valgus knee treated with CWDFO at Department of Integrated Chinese and Western Medicine Orthopedics, Honghui Hospital, Xi'an Jiaotong University from January 2018 to July 2019 were analyzed retrospectively. There were 3 males and 11 females, aging (41.4±16.4) years (range: 18 to 56 years). The body mass index was (23.5±3.5) kg/m(2) (range: 18.1 to 28.9 kg/m(2)). The guide pins were placed with the assistance of the self-designed AAOG. Before the surgery, Solidworks software was used to calculate the correction angle and the osteotomy radius accurately. The osteotomy guide was adjusted according to these two parameters. During the surgery, the adjusted osteotomy guide was placed to the surface of bone closely and the guide pins were drilled into the bone through the guide holes. The position of the guide pins was confirmed under fluoroscopy. The osteotomy was finished under guide of pins and fixed with Tomofix plate (Synthes). The times and duration of placement of the guide pins, the times of X-ray examination, the planned and actual thickness of the osteotomy wedge, the top and bottom area of the osteotomy wedge, the posterior distal femoral angle(PDFA), the correction of the weight line, and the American Knee Society Score(AKSS) and Tegner scores were collected and compared by paired t test or Kruskal-Wallis non-parametric test. Healing time after osteotomy and complications were recorded. Results: The guide pins were successfully placed once in 10 knees, adjusted once in 5 knees and twice in 2 knees. The time spent in placing all the 6 pins was 82.4 seconds (range: 51 to 125 seconds), and the times of X-ray examination was 1.5 times (range: 1 to 5 times). The top and bottom areas of the osteotomy wedge were (5.52±0.52)cm(2) and (5.36±0.49)cm(2). PDFA was (85.2±2.6)° preoperatively and (85.5±1.4)° postoperatively (t=-0.401, P>0.05). The thickness of the osteotomy was (11.3±1.9)mm according to the preoperative plan, and the actual thickness was (8.1±1.7)mm. All the patients were followed up for 6 months after surgery and AKSS and Tegner scores improved significantly (all P<0.05). The correction of the weight lines was within the ideal range. Fractures of the hinge point occurred in 3 knees. All of the osseous healing without complications. Conclusion: The new osteotomy guide helps to place the guide pins rapidly and precisely according to the preoperative planning, which should be widely used in clinical applications with promising outcomes.

[Efficacy and peripheral immunity analysis of allogeneic natural killer cells therapy in patients with hepatocellular carcinoma].

OBJECTIVE: To evaluate the safety and efficacy of allogeneic natural killer (NK) cells in the treatment of primary hepatocellular carcinoma (HCC), and to elucidate the mechanism of NK cells therapy. METHODS: Twenty-one patients with primary HCC treated with allogeneic NK cells at the Fifth Medical Center of the PLA General Hospital were followed up for 1 year. Peripheral blood mononuclear cells (PBMCs) were isolated from patient-related donors and cultured in vitro for 15 days and infused to the patients in two consecutive days. Clinical data and laboratory data were collected and analyzed, including survival, clinical features, imaging changes, hematology, immunology, and biochemical indicators to evaluate the safety and efficacy of allogeneic NK cell therapy. The changes of peripheral blood lymphocyte subsets after treatment were also analyzed to explore the possible anti-tumor mechanisms. RESULTS: (1) Of the 21 patients with primary HCC, 11 patients were treated once, 5 patients were treated twice, and 5 patients were treated 3 times. After allogeneic NK cells infusion, 10 patients had fever, 1 patient had slight hepatalgia and 1 patient had slight headache, no other adverse events occurred including acute and chronic graft-versus-host disease (GVHD). They resolved spontaneously within 8 hours without other treatment. (2) The total disease control rate was 76.2% during one-year follow-up. Among them, the patients with Barcelona clinic liver cancer (BCLC) stage A had a disease control rate of 100%, stable disease (SD) in 10 cases; BCLC stage B patients had a disease control rate of 60%, partial response (PR) in 1 case, and SD 2 in cases; BCLC stage C patients had a disease control rate of 50%, complete response (CR) in 1 case, and 2 cases of PR. (3) The frequencies of NK cells and CD8+ T cells in peripheral blood were significantly lower than that before at 24 hours after treatment, and the frequencies of CD4+ T cells and CD4/CD8 were significantly higher than the baseline. CONCLUSION: Allogeneic NK cells have good safety and efficacy in the treatment of primary HCC. The anti-tumor effect of the allogeneic NK cells may play an important role in the activation of the patient's natural immune system and delay disease progression, suggesting that allogeneic NK cells combined with sorafenib may be a very effective treatment for advanced HCC, and further large-sample multicenter randomized controlled clinical trials are needed to validate this result.

Screening and analysis on the differentially expression genes between diploid and autotetraploid watermelon by using of digital gene expression profile / Triagem e análise da expressão diferencial dos genes entre melancia diplóide e autotetraplóide através do perfil de expressão gênica digital

Abstract Synthetic polyploids are key breeding materials for watermelon. Compared with diploid watermelon, the tetraploid watermelon often exhibit wide phenotypic differences and differential gene expression. Digital gene expression (DGE) profile technique was performed in this study to present gene expression patterns in an autotetraploid and its progenitor diploid watermelon, and deferentially expressed genes (DEGs) related to the abiotic and biotic stress were also addressed. Altogether, 4,985 DEGs were obtained in the autotetraploid against its progenitor diploid, and 66.02% DEGs is up-regulated. GO analysis shows that these DEGs mainly distributed in 'metabolic process', 'cell' and 'catalytic activity'. KEGG analysis revealed that these DEGs mainly cover 'metabolic pathways', 'secondary metabolites' and 'ribosome'. Moreover, 134 tolerance related DEGs were identified which cover osmotic adjustment substance, protective enzymes/protein, signaling proteins and pathogenesis-related proteins. This study present the differential expression of stress related genes and global gene expression patterns at background level in autotetraploid watermelons. These new evidences could supplement the molecular theoretical basis for the better resistance after the genome doubling in the gourd family.

Resumo Poliploides sintéticos são materias fundamentais para melhoramento genético da melancia. Comparativamente ao seu homólogo diploide, a melancia tetraploide apresenta amplas diferenças genotípica e fenotípica e diferença de expressão gênica. A expressão gênica digital ou DGE (digital gene expression) foi utilizada neste estudo para representar o perfil de expressão gênica da melancia autotetraploide e seu progenitor diploide e a expressão diferencial de genes relacionados ao estresse biótico e abiótico. Os resultados mostraram que 4.985 DEGs foram observados no organismo autotetraploide, sendo que, deste total, 66.02%foram supra-regulados. A análise de ontologia gênica (GO) mostrou que estes DEGs estão relacionados principalmente com processos metabólicas, célula e atividade catalítica, abrangendo de acordo com a análise de genes e genoma (KEGG) rotas metabólicas, metabolismo secundário e ribossomos. Além disso, 134 genes de defesa foram identificados, abrangendo substâncias de ajuste osmótico, enzimas/proteínas de proteção, proteínas sinalizadoras e proteínas relacionadas à patogênese. Este estudo mostrou a expressão diferencial de genes relacionados ao estresse e o perfil global de expressão gênica de melancia autotetraploide, estes resultados podem complementar, a nível molecular, o entendimento do fator resistência após a duplicação do genoma em cucurbitáceas.

Screening and analysis on the differentially expression genes between diploid and autotetraploid watermelon by using of digital gene expression profile.

Synthetic polyploids are key breeding materials for watermelon. Compared with diploid watermelon, the tetraploid watermelon often exhibit wide phenotypic differences and differential gene expression. Digital gene expression (DGE) profile technique was performed in this study to present gene expression patterns in an autotetraploid and its progenitor diploid watermelon, and deferentially expressed genes (DEGs) related to the abiotic and biotic stress were also addressed. Altogether, 4,985 DEGs were obtained in the autotetraploid against its progenitor diploid, and 66.02% DEGs is up-regulated. GO analysis shows that these DEGs mainly distributed in 'metabolic process', 'cell' and 'catalytic activity'. KEGG analysis revealed that these DEGs mainly cover 'metabolic pathways', 'secondary metabolites' and 'ribosome'. Moreover, 134 tolerance related DEGs were identified which cover osmotic adjustment substance, protective enzymes/protein, signaling proteins and pathogenesis-related proteins. This study present the differential expression of stress related genes and global gene expression patterns at background level in autotetraploid watermelons. These new evidences could supplement the molecular theoretical basis for the better resistance after the genome doubling in the gourd family.

Screening and analysis on the differentially expression genes between diploid and autotetraploid watermelon by using of digital gene expression profile

Abstract Synthetic polyploids are key breeding materials for watermelon. Compared with diploid watermelon, the tetraploid watermelon often exhibit wide phenotypic differences and differential gene expression. Digital gene expression (DGE) profile technique was performed in this study to present gene expression patterns in an autotetraploid and its progenitor diploid watermelon, and deferentially expressed genes (DEGs) related to the abiotic and biotic stress were also addressed. Altogether, 4,985 DEGs were obtained in the autotetraploid against its progenitor diploid, and 66.02% DEGs is up-regulated. GO analysis shows that these DEGs mainly distributed in metabolic process, cell and catalytic activity. KEGG analysis revealed that these DEGs mainly cover metabolic pathways, secondary metabolites and ribosome. Moreover, 134 tolerance related DEGs were identified which cover osmotic adjustment substance, protective enzymes/protein, signaling proteins and pathogenesis-related proteins. This study present the differential expression of stress related genes and global gene expression patterns at background level in autotetraploid watermelons. These new evidences could supplement the molecular theoretical basis for the better resistance after the genome doubling in the gourd family.

Resumo Poliploides sintéticos são materias fundamentais para melhoramento genético da melancia. Comparativamente ao seu homólogo diploide, a melancia tetraploide apresenta amplas diferenças genotípica e fenotípica e diferença de expressão gênica. A expressão gênica digital ou DGE (digital gene expression) foi utilizada neste estudo para representar o perfil de expressão gênica da melancia autotetraploide e seu progenitor diploide e a expressão diferencial de genes relacionados ao estresse biótico e abiótico. Os resultados mostraram que 4.985 DEGs foram observados no organismo autotetraploide, sendo que, deste total, 66.02%foram supra-regulados. A análise de ontologia gênica (GO) mostrou que estes DEGs estão relacionados principalmente com processos metabólicas, célula e atividade catalítica, abrangendo de acordo com a análise de genes e genoma (KEGG) rotas metabólicas, metabolismo secundário e ribossomos. Além disso, 134 genes de defesa foram identificados, abrangendo substâncias de ajuste osmótico, enzimas/proteínas de proteção, proteínas sinalizadoras e proteínas relacionadas à patogênese. Este estudo mostrou a expressão diferencial de genes relacionados ao estresse e o perfil global de expressão gênica de melancia autotetraploide, estes resultados podem complementar, a nível molecular, o entendimento do fator resistência após a duplicação do genoma em cucurbitáceas.

[Multiposition spiral suture of the lower uterine segment: a new technique to control the intraoperative bleeding of pernicious placenta previa].

Objective: To explore the efficacy and safety of multiposition spiral suture of the lower uterine segment, a new technique to control the intraoperative bleeding of pernicious placenta previa(PPP). Methods: From May 2014 to May 2015, 38 patients were diagnosed PPP in Tongji Hospital and cesarean sections were performed. After removing the placenta, multiposition spiral suture was used when massive bleeding occurred, and bilateral descending branches of uterine artery ligation was conducted when necessary. Results: 18 of the 38 PPP patients(47%,18/38)were diagnosed placenta accreta. The average cervical canal length of 38 PPP patients was(3.1±0.6)cm. There were 12 cases(32%, 12/38)with 4 regions sutured, 23 cases(61%, 23/38)with 2-3 regions sutured and 3 cases(8%, 3/38)with only posterior wall area sutured. Twelve cases(32%, 12/38)underwent uterine artery ligation, 3 cases(8%, 3/38)underwent uterine artery ligation and COOK balloon. None of them was postpartum hemorrhage or performing internal iliac artery embolization. Two patients received hysterectomy. The average blood loss in the operation was(1 696± 1 397)ml. In 16(42%,16/38)patients, the blood loss exceeded 1 500 ml, and the heaviest one was 4 500 ml. Three patients had haematuria in the first 3 days after the operation. No complication was found in 6 months after the operation. Conclusions: The multiposition spiral suture technique is a simple, safe and effective way to control the massive bleeding in the cesarean section of PPP patients. It is also beneficial for the recovery of the uterus.

[Influences of inhaling glucocorticoid on sIgA and HBD-2,3 in pharyngeal mucosa of and T cell subsets in blood of patients with chronic persistent asthma].

Objective:The aim of this study was to investigate the inhaled glucocorticoid therapy affects sIgA and HBD 2,3 secreted by mucous membrane of throat and T lymphocyte subsets in patients with chronic persistent asthma. Method:One hundred patients diagnosed with chronic persistent asthma were randomly divided into control group (submitted to conventional non-hormonal therapy) and treatment group (submitted to inhaled glucocorticoid therapy) with 50 cases in each group. Ventolin aerosol had been promised in on demand use and the dosage of nonhormonal drugs should be adjusted according to the severity of illness in each group. No matter if asthma was fully under control or not, the observation would be terminated in 3 months. The secretion samples of pharyngeal mucosa and venous blood samples had been collected before and after treatment. The content of sIgA, HBD-2 and HBD-3 were detected by ELISA method. T lymphocyte subsets were detected by flow cytometry.Result:Compared with control group, the content of sIgA, HBD-2 and HBD-3 were lower in treatment group(P<0.05). Nevertheless, there were no significant differences of T lymphocyte subsets including CD3+,CD4+ and CD4+/CD8+ between two groups(P>0.05).Conclusion:The inhaled glucocorticoid therapy could bring adverse effects on the immune function of local mucous membrane in patients with asthma, with no significant effect on peripheral blood T cell subsets.

Preperitoneal femoral hernioplasty: an "umbrella" technique.

INTRODUCTION: Tension-free inguinal mesh-plug hernioplasty is well established. However, femoral hernia repair remains challenging and controversial. We aimed to evaluate a preperitoneal approach of tension-free hernioplasty for femoral hernia upon the anatomy rationality. METHODS: A prospective study of 62 patients between October 1999 and June 2011 received femoral hernioplasty in our hospital. This repair method involved a preperitoneal approach accomplished under regional or local anesthesia with mesh and plug; the emphasis was put on fulfilling the abdominal defect, i.e., the myopectineal orifice, with the plug flattened like an "umbrella", above the femoral ring but not to fill the femoral ring. RESULTS: All cases receiving preperitoneal tension-free hernioplasty had a smooth recovery. There were no severe complications, and no recurrences were detected within a 0.5- to 4-year follow-up. No specific restrictions with regard to activity were placed on the patients after surgery. All cases were able to return to normal life, including work, within 2 weeks. CONCLUSIONS: The preperitoneal tension-free hernioplasty may be a more effective method of femoral hernia repair; meanwhile, we must re-understand the anatomy of femoral hernia correctly so as to restore the anatomic and physiologic functions at this region optimally.

Promoter hypomethylation and increased maspin expression in preeclamptic placentas in a Chinese population.

OBJECTIVE: Preeclampsia is thought to begin with shallow trophoblast invasion and inadequate spiral artery remodeling. Maspin, a tumor-suppressor gene, plays a regulatory role in trophoblast invasion and motility. The tissue-specific methylation of the maspin promoter can regulate maspin gene expression in various cancers. We sought to detect maspin gene expression and assess the degrees of methylation of maspin promoter regions in preeclamptic placentas in the Han Chinese population and to investigate the potential role of maspin in the pathophysiology of preeclampsia. METHODS: We conducted RT-PCR, immunohistochemistry and western blotting to characterize maspin gene expression and protein levels in the placentas from normal and preeclamptic pregnancies. Finally, using methylation-specific PCR and bisulfite sequencing PCR, we detected the degrees of methylation of the promoter regions of maspin in each of the two studied groups. RESULTS: Maspin expression was increased at the mRNA and protein levels in the preeclamptic placentas compared to the control group. Maspin immunohistochemical staining revealed positive staining in the syncytio-cytotrophoblast layers and more diffuse staining in the preeclamptic group. The mean methylation level of the analyzed promoter region was significantly hypomethylated in the preeclamptic placentas compared to the control placentas, pointing to a negative relationship between maspin promoter methylation and gene expression. DISCUSSION: Hypomethylation of the maspin promoter results in increased expression of maspin in preeclamptic placentas, which suggests a negative relationship between maspin methylation and maspin expression in this Han Chinese population. Thus, maspin is likely involved in the etiology of preeclampsia.

A novel protocol allowing oral delivery of a protein complement inhibitor that subsequently targets to inflamed colon mucosa and ameliorates murine colitis.

While there is evidence of a pathogenic role for complement in inflammatory bowel disease, there is also evidence for a protective role that relates to host defence and protection from endotoxaemia. There is thus concern regarding the use of systemic complement inhibition as a therapeutic strategy. Local delivery of a complement inhibitor to the colon by oral administration would ameliorate such concerns, but while formulations exist for oral delivery of low molecular weight drugs to the colon, they have not been used successfully for oral delivery of proteins. We describe a novel pellet formulation consisting of cross-linked dextran coated with an acrylic co-polymer that protects the complement inhibitor CR2-Crry from destruction in the gastrointestinal tract. CR2-Crry containing pellets administered by gavage, were characterized using a therapeutic protocol in a mouse model of dextran sulphate sodium (DSS)-induced colitis. Oral treatment of established colitis over a 5-day period significantly reduced mucosal inflammation and injury, with similar therapeutic benefit whether or not the proton pump inhibitor, omeprazole, was co-administered. Reduction in injury was associated with the targeting of CR2-Crry to the mucosal surface and reduced local complement activation. Treatment had no effect on systemic complement activity. This novel method for oral delivery of a targeted protein complement inhibitor will reduce systemic effects, thereby decreasing the risk of opportunistic infection, as well as lowering the required dose and treatment cost and improving patient compliance. Furthermore, the novel delivery system described here may provide similar benefits for administration of other protein-based drugs, such as anti-tumour necrosis factor-α antibodies.

Lixisenatide rescues spatial memory and synaptic plasticity from amyloid ß protein-induced impairments in rats.

Alzheimer's disease (AD) is a progressive and degenerative disorder accompanied by cognitive impairment, but effective strategies against AD are currently not available. Interestingly, glucagon-like peptide-1 (GLP-1) used in type 2 diabetes mellitus (T2DM) has shown neuroprotective effects in preclinical studies of AD. Lixisenatide, an effective GLP-1 receptor (GLP-1R) agonist with much longer half life than GLP-1, has been licensed in the EU as a treatment for T2DM. However, the neuroprotective effects of lixisenatide in the brain remain to be clarified. In the present study, we report for the first time the effects of lixisenatide on the amyloid ß (Aß) protein-induced impairments in spatial learning and memory of rats, and investigated its electrophysiological and molecular mechanisms. We found that: (1) bilateral intrahippocampal injection of Aß25-35 resulted in a significant decline in spatial learning and memory of rats, as well as a suppression of in vivo hippocampal long-term potentiation (LTP); (2) lixisenatide treatment effectively prevented the Aß25-35-induced impairments; (3) lixisenatide inhibited the Aß25-35 injection-induced activation of glycogen synthase kinase 3ß (GSK3ß), with a significant increase in the phosphorylation of ser9 and a significant decrease in the phosphorylation of Y216. These results indicate that lixisenatide, by affecting the PI3K-Akt-GSK3ß pathway, can prevent Aß-related impairments in synaptic plasticity and spatial memory of rats, suggesting that lixisenatide may be a novel and effective treatment for AD.

HBx-mediated miR-21 upregulation represses tumor-suppressor function of PDCD4 in hepatocellular carcinoma.

The hepatitis B virus (HBV) X protein (HBx) has a key role in the molecular pathogenesis of HBV-related hepatocellular carcinoma (HCC). However, the mechanism of HBx-mediated hepatocarcinogenesis remains to be elucidated. In this study, we aimed to better understand the effects of HBx on gene-expression profiles that participate in hepatocarcinogenesis and the mechanism by which HBx regulates these genes. Differentially expressed genes between L02-HBx and L02-Vector control cells were identified by microarray and validated using quantitative real-time PCR. HBx upregulates 456 genes and downregulates 843 genes, including programmed cell death 4 (PDCD4). PDCD4 was downregulated in clinical HCC specimens and the downregulation of PDCD4 in HCC is correlated with HBx. Furthermore, overexpression experiments in HCC cells proved that PDCD4 has strong tumor-suppressive effects both in vitro and in vivo, and may induce cell apoptosis to suppress the development of HCC. HBx induces expression of DNA methyltransferases (DNMTs), but failed to change the methylation status of the PDCD4 promoter. HBx downregulates PDCD4 expression at least partially through miR-21. Taken together, this study reported for the first time that HBx downregulates PDCD4 and upregulates miR-21 expression. The overexpression of PDCD4 could suppress tumorigenicity. The deregulation of PDCD4 by HBx through miR-21 represents a potential novel mechanism of the downregulation of PDCD4 in HBV-related HCC and provides new insights into HCC development.

Cdk5 targets active Src for ubiquitin-dependent degradation by phosphorylating Src(S75).

The non-receptor tyrosine kinase Src is a critical regulator of cytoskeletal contraction, cell adhesion, and migration. In normal cells, Src activity is stringently controlled by Csk-dependent phosphorylation of Src(Y530), and by Cullin-5-dependent ubiquitinylation, which affects active Src(pY419) exclusively, leading to its degradation by the proteosome. Previous work has shown that Src activity is also limited by Cdk5, a proline-directed kinase, which has been shown to phosphorylate Src(S75). Here we show that this phosphorylation promotes the ubiquitin-dependent degradation of Src, thus restricting the availability of active Src. We demonstrate that Src(S75) phosphorylation occurs in vivo in epithelial cells, and like ubiquitinylation, is associated only with active Src. Preventing Cdk5-dependent phosphorylation of Src(S75), by site-specific mutation of S75 or by Cdk5 inhibition or suppression, increases Src(Y419) phosphorylation and kinase activity, resulting in Src-dependent cytoskeletal changes. In transfected cells, ubiquitinylation of Src(S75A) is about 35% that of wild-type Src-V5, and its half-life is approximately 2.5-fold greater. Cdk5 suppression leads to a comparable decrease in the ubiquitinylation of endogenous Src and a similar increase in Src stability. Together, these findings demonstrate that Cdk5-dependent phosphorylation of Src(S75) is a physiologically significant mechanism of regulating intracellular Src activity.

Lamivudine in late pregnancy to prevent perinatal transmission of hepatitis B virus infection: a multicentre, randomized, double-blind, placebo-controlled study.

This randomized, double-blind, placebo-controlled study evaluated whether lamivudine given during late pregnancy can reduce hepatitis B virus (HBV) perinatal transmission in highly viraemic mothers. Mothers were randomized to either lamivudine 100 mg or placebo from week 32 of gestation to week 4 postpartum. At birth, infants received recombinant HBV vaccine with or without HBIg and were followed until week 52. One hundred and fifty mothers, with a gestational age of 26-30 weeks and serum HBV DNA >1000 MEq/mL (bDNA assay), were treated. A total of 141 infants received immunoprophylaxis at birth. In lamivudine-treated mothers, 56 infants received vaccine + HBIg (lamivudine + vaccine + HBIg) and 26 infants received vaccine (lamivudine + vaccine). In placebo-treated mothers, 59 infants received vaccine + HBIg (placebo + vaccine + HBIg). At week 52, in the primary analyses where missing data was counted as failures, infants in the lamivudine + vaccine + HBIg group had a significant decrease in incidence of HBsAg seropositivity (10/56, 18%vs 23/59, 39%; P = 0.014) and in detectable HBV DNA (11/56, 20%vs 27/59, 46%; P = 0.003) compared to infants in the placebo + vaccine + HBIg group. Sensitivity analyses to evaluate the impact of missing data at week 52 resulting from a high dropout rate (13% in the lamivudine + vaccine + HBIg group and 31% in the placebo + vaccine + HBIg group) remained consistent with the primary analysis in that lower transmission rates were still observed in the infants of lamivudine-treated mothers, but the differences were not statistically significant. No safety concerns were noted in the lamivudine-treated mothers or their infants. Results of this study suggest that lamivudine reduced HBV transmission from highly viraemic mothers to their infants who received passive/active immunization.

Fanconi anemia proteins localize to chromatin and the nuclear matrix in a DNA damage- and cell cycle-regulated manner.

Fanconi anemia (FA) is a genetic disease characterized by congenital defects, bone marrow failure, and cancer susceptibility. Cells from patients with FA exhibit genomic instability and hypersensitivity to DNA cross linking agents such as mitomycin C. Despite the identification of seven complementation groups and the cloning of six genes, the function of the encoded gene products remains elusive. The FancA (Fanconi anemia complementation group A), FancC, and FancG proteins have been detected within a nuclear complex, but no change in level, binding, or localization has been reported as a result of drug treatment or cell cycle. We show that in immunofluorescence studies, FancA appears as a non-nucleolar nuclear protein that is excluded from condensed, mitotic chromosomes. Biochemical fractionation reveals that the FA proteins are found in nuclear matrix and chromatin and that treatment with mitomycin C results in increase of the FA proteins in nuclear matrix and chromatin fractions. This induction occurs in wild-type cells and mutant FA-D (Fanconi complementation group D) cells but not in mutant FA-A cells. Immunoprecipitation of FancA protein in chromatin demonstrates the coprecipitation of FancA, FancC, and FancG, showing that the FA proteins move together as a complex. Also, fractionation of mitotic cells confirms the lack of FA proteins in chromatin or the nuclear matrix. Furthermore, phosphorylation of FancG was found to be temporally correlated with exit of the FA complex from chromosomes at mitosis. Taken together, these findings suggest a role for FA proteins in chromatin and nuclear matrix.

Human lens thioltransferase: cloning, purification, and function.

PURPOSE: To clone the human lens thioltransferase (TTase) gene and to purify, characterize and study the possible function of the recombinant human lens thioltransferase (RHLT). METHODS: The human lens TTase gene was cloned by using RT-PCR and verified by sequence and RNase protection assay. TTase overexpressed in Escherichia coli was isolated and purified to homogeneity by column chromatography and identified by Western blot analysis. The activity was assayed with a synthetic substrate hydroxyethyl disulfide. Its function in dethiolating and reactivating other key metabolic enzymes was studied by using pure glutathione S:-transferase (GST) and glutathione peroxidase (GPx) from commercial source and also with the cell extract of rabbit lens epithelial cells preexposed to H2O2. RESULTS: The cloned human lens TTase gene showed identical sequence to the TTase gene from other human tissues. The RNase protection assay displayed a single transcript from the total RNA of human lens epithelial cells. The purified RHLT had a molecular weight of 11.8 kDa and reacted positively with anti-pig liver TTase. It displayed similar structural, functional, and kinetic characteristics to those of TTases from other sources. It was shown that RHLT effectively regenerated the activities of GST and GPx, after each was inactivated by S-thiolation with cystine in vitro. Furthermore, RHLT was able to restore the activity of the oxidatively inactivated glyceraldehyde-3-phosphate dehydrogenase (G-3PD) in H2O2-exposed rabbit lens epithelial cells. CONCLUSIONS: The human lens TTase gene has been cloned for the first time. Its gene product showed the characteristics which support our speculation that TTase may play a major role in maintaining the homeostasis of lens protein thiols thus protecting against oxidative stress.