M2e gene of three copies for H5N1 subtype AIV was synthesized and fused with human mycobacterium tuberculosis hsp70 gene.The fused gene was cloned into the prokaryotic expression vector to get pET-3M2e and pET-3M2e-hsp70.Recombinant protein r3M2e and r3M2e-hsp70 were successfully expressed induced with IPTG and purified with Ni2+-NTA collumn.Following that,the immunity of the recombinant protein was analysized with Western blot.20-day-old AIV non-immunized chickens were vaccined with r3M2e and r3M2e hsp70,at the same time,Trx and KLH-M2e inoculated chickens were served as vector and positive controls.Two weeks after the primary vaccination,every group was boosted with the same vaccine as in the primary vaccination.The humoral immunity of the vaccined chickens was evaluated with antibody detection against M2e,cytopathic suppression test,and indirect fluorescence assay.The cellular immunity was estimated according to lymphocyte subtype analysis with flow cytometry and M2e specific cytokine detection.Four weeks after the boost vaccine,all groups were challenged with 100EID50 AIV of H9N2 subtype,and the virus from swabs was detected with Real-time PCR.Results indicated that r3M2e hsp70 vaccined chicken developed the better humoral and cellular immune response,also,made a better performance compared with r3M2e vaccined group in virus challenge.

Objective To investigate CT and MRI features of dermatofibrosarcoma protuberans (DFSP).Methods Totally 16 patients with DFSP confirmed by pathology were enrolled.Tumor morphology,CT and MRI imaging appearance (11 cases underwent plain and enhanced CT,5 cases underwent plain and enhanced MRI) were analyzed retrospectively.Results DFSP usually occurred in the skin of truck,head and neck,protruding from the skin surface in different extent.Some lesions even suspended out of the skin.The lesions were divided into nodular type (n=12) and diffuse type (n=4) according to their morphological appearance.The tumors usually demonstrated as iso-density or slightly low density solid mass compared to muscle on CT.On MRI,it usually demonstrated as low signal on T1WI and high signal on T2WI.Tumor blood supply was rich,and it usually showed progressively moderate to strong enhancement.The signs within DFSP include hanging sign (n=2),skin tail sign (n＝6),fascia tail sign (n=l),fat tail sign (n=4).Conclusion DFSP can be characterized by nodular or diffuse lesions,the manifestations of different form are slightly different,but still have a certain characteristic.

BACKGROUND:The phenomenon of osteoinduction by biomaterials has been proven in animal experiments. OBJECTIVE:To investigate whether the ability of a biomaterial to initiate bone formation in ectopic implantation sites improves the performance of osteoinductive biomaterial as a scaffold for tissue-engineered bone. METHODS:We compared ectopic bone formation by combining autologous adipose-derived stromal cells with an osteoinductive and a nonosteoinductive biphasic calcium phosphate ceramic to create a tissue engineering construction in the muscle of dogs. Al implants were implanted in the back muscle of 10 adult dogs for 8 weeks and 12 weeks, including osteoinductive biphasic calcium phosphate ceramic+adipose-derived stromal cells (osteoinductive complex group), osteoinductive biphasic calcium phosphate ceramic (osteoinductive broup), nonosteoinductive biphasic calcium phosphate ceramic+adipose-derived stromal cells (nonosteoinductive complex group), and nonosteoinductive biphasic calcium phosphate ceramic (nonosteoinductive group). Micro-CT analysis and histomorphometry were performed to evaluate and quantify ectopic bone formation. RESULTS AND CONCLUSION:Ectopic bone formation was visible in the osteoinductive complex group and osteoinductive group, and the former group was superior to the latter one in quality of new bone (P<0.05). However, there was no ectopic bone formation in the other two groups. Micro-CT results were consistent with the histomorphological detection. These findings indicate that osteoinductive biphasic calcium phosphate ceramic, as a kind of bone tissue engineering scaffold material, has a better osteogenic capacity, while adipose-derived mesenchymal stem cells serve as seed cells to promote the ectopic bone formation.

In this study, a composite cell model for evaluation of idiosyncratic drug-induced liver injury (IDILI) was established in vitro from the perspective of immune inflammation. And this model was used to evaluate the risk of IDILI for 2,3,5,4'-tetrahydroxy-cis-stilbene-2-O-β-glucoside (Cis-SG) and 2,3,5,4'-tetrahydroxy-trans-stilbene-2-O-β-glucoside (Trans-SG). To determine the low, medium, and high dosage of Cis-SG and Trans-SG, CellTiter-Glo^® 3D Cell Viability Assay was used to detect the effects of Cis-SG and Trans-SG on cell viability of HepG2 cells in three dimensional (3D) culture, and MTT assay was used to detect the effects of Cis-SG and Trans-SG on cell viability of THP-1 derived macrophages. THP-1 derived macrophages were incubated by Cis-SG and Trans-SG directly or supernatants from HepG2 cells incubated with them. Enzyme linked immunosorbent assay (ELISA) was used to detect the levels of interleukin-1β (IL-1β) in the supernatants of the THP-1 derived macrophages. Western blot and reverse transcription-polymerase chain reaction (RT-PCR) were used to determine the expression of apoptosis-associated speck-like protein (ASC), Nod-like receptor protein 3 (NLRP3), cysteinyl aspartate specific proteinase-1 (caspase-1), and IL-1β in THP-1 derived macrophages. The results showed that there was no effect on the secretion of IL-1β in THP-1 derived macrophages incubated by Cis-SG and Trans-SG directly. However, the secretion of IL-1β, the protein and mRNA expression of ASC, NLRP3, caspase-1, and IL-1β significantly increased in THP-1 derived macrophages incubated by supernatants from HepG2 cells incubated with 1, 5, and 25 μmol·L^-1 Cis-SG or 25 μmol·L^-1 Trans-SG. In summary, the composite cell model for evaluation of IDILI established in vitro has been successfully applied in testing Cis-SG and Trans-SG. This composite cell model is helpful to evaluate and screen drugs with IDILI risk in vitro preliminarily, which provides methods for predicting and solving the idiosyncratic liver toxicity of drugs.

Objective To study interleukin-23 (IL-23) levels in serum and dendritic cells of acute-on-chronic liver failure (ACLF) patients with chronic hepatitis B (CHB) and to explore its relationship with the prognosis.Methods Peripheral blood mononuclear cells and serum were collected from 40 ACLF patients with CHB (including survival group 27 cases and non-survival group 13 cases) and 26 healthy controls.Monocytes were induced to immature dendritic cell in vitro and TNF-α was added to induce dendritic cell maturation.IL-23 mRNA of dendritic cells was detected by real time polymerase chain reaction (PCR), and serum IL-23 level was measured by enzyme-linked immuno sorbent assay (ELISA).Differences among the parameters with normal distribution were compared using t test, those with non-normal distribution were compared using non-parametric Mann-Whitney U-test, and the relationship between two variables was assessed by Spearman′s rank correlation.Results International normalized rate (INR) and model for end-stage liver disense (MELD) scores in non-survival group of ACLF were higher than those in survival group (INR: 2.32 vs 1.64, U=69.00, P=0.002 2;MELD:36 vs 30, U=64.50, P=0.001 4).However, there were no significant differences between two groups at gender, age, alanin aminotransferase (ALT), aspartate aminotrans ferase (AST), bilirubin, creatinine, hepatitis B virus (HBV) DNA, hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg) and serum IL-23.IL-23 mRNA level in dendritic cells at baseline in non-survival group of ACLF was significantly higher than that in survival group (76 vs 43, U=71.50, P=0.002 8).After treatment, serum IL-23 was significantly declined in survival group ([160±75] ng/L vs [91±49] ng/L, t=4.012, P=0.000 2), but not in non-survival group.Significant positive correlation was observed between IL-23 mRNA level in dendritic cells and MELD score at baseline (r=0.7198,P<0.01).Conclusions Persistent high serum IL-23 level suggests poor prognosis in ACLF patients with CHB.IL-23 mRNA expression in dendritic cells has good consistency with MELD score and the patients with high IL-23 mRNA expression has poor outcome.

OBJECTIVETo assess clinical therapeutic effect and safety of acupuncture combined with fuxiting on depression.

METHODSFifty-three cases of depression were divided into an observation group (n = 30) treated by acupuncture combined with fuxiting and a control group treated by fuxiting. Their clinical therapeutic effects were assessed by HAMD, HAMA and CGI, and adverse effects by TESS.

RESULTSAfter treatment of 6 weeks, the total effective rate was 80.0% in the observation group and 69.6% in the control group with no significant difference between the two groups (P > 0.05). There were significant differences in scores of HAMD and HAMA before and after treatment in the two groups (P < 0.01), with no significant difference between the two groups (P > 0.05). The adverse effect in the observation group was less and milder than that in the control group.

CONCLUSIONAcupuncture combined fuxiting has good therapeutic effect on depression with less and mild adverse effects.

Acupuncture Therapy ; Combined Modality Therapy ; Depression ; therapy ; Depressive Disorder ; Humans

Biopsy ; Endopeptidase K ; Fluorescent Antibody Technique ; Glomerulonephritis, IGA ; pathology ; Glomerulonephritis, Membranous ; pathology ; Humans ; Kidney ; pathology ; Lupus Nephritis ; pathology ; Paraffin Embedding ; Staining and Labeling

Objective:To summarize the characteristics of genetic variation and prenatal diagnosis in pedigrees with X-linked adrenoleukodystrophy (X-ALD) and elucidate the value of prenatal diagnosis in preventing the birth of children with X-ALD.Methods:Twenty pedigrees, clinically diagnosed with X-ALD in Peking University First Hospital from November 2012 and March 2019, were included in this retrospective study. Genomic DNA was extracted from peripheral blood and amniotic fluid or chorionic villi samples of probands and their families for detecting variants in ATP-binding cassette subfamily D member 1 ( ABCD1) gene using polymerase chain reaction (PCR)-Sanger sequencing. Linkage analysis was also performed on five microsatellite markers near ABCD1 gene to exclude maternal contamination. Characteristics of ABCD1 gene variants and prenatal diagnosis of X-ALD pedigrees were summarized by descriptive statistics. Results:Twenty ABCD1 gene variants were identified in the 20 pedigrees. The variants in three probands that were not detected by next-generation sequencing were identified by PCR-Sanger sequencing. Among the mothers of the 20 probands, 17 carried ABCD1 variants and three did not. We performed 24 prenatal diagnoses on 20 pregnancies (24 fetuses) and identified eight fetuses with variants who were finally terminated. The 16 cases without variants were born alive. The validation results obtained after termination or delivery were consistent with those performed prenatally. Conclusions:No hotspot variants in ABCD1 gene are detected in these X-ALD patients and most variants are maternally inherited. PCR-Sanger sequencing is an effective method for detecting ABCD1 variants. Prenatal diagnosis for mothers who had a body with X-ALD could prevent another one from birth.

OBJECTIVETo detect the IgH-MMSET fusion gene resulted from t (4;14) translocation in multiple myeloma and illuminate its significance.

METHODSIgH-MMSET fusion gene was detected in bone marrow specimens of 25 multiple myeloma (MM) patients and MM cell line NCI-H929 using reverse-transcription PCR (RT-PCR) assay followed by nested PCR to increase the sensitivity. The purified PCR products were cloned into pGEM-T vector and then sequenced using M13 forward primers. The fragment sequences were compared with that in GenBank to find matched sequences.

RESULTSOnly a 438 base pair long fragment was obtained after RT-PCR assay and was confirmed by sequencing to be a fusion gene product of IgH gene and MMSET gene in MM cell line NCI-H929. The breakpoints were located within the C micro region of IgH gene on chromosome 14 and intron 3 of MMSET gene on chromosome 4. IgH-MMSET hybrid transcripts were detected in 3 of 25 MM patients through nested PCR assay. The amplified fragments of the 3 patients were 237 base pairs (bp), 239 bp and 239 bp in length, respectively. The breakpoints on chromosome 4 were identical to that of NCI-H929 cell.

CONCLUSIONSThe formation of IgH-MMSET fusion gene is resulted from t (4;14) translocation in MM. The incidence rate is 12.0%. The presence of IgH-MMSET fusion gene may predict poor prognosis.

Adult ; Aged ; Base Sequence ; Chromosomes, Human, Pair 14 ; Chromosomes, Human, Pair 4 ; Female ; Humans ; Male ; Middle Aged ; Molecular Sequence Data ; Multiple Myeloma ; genetics ; Oncogene Proteins, Fusion ; genetics ; Protein-Tyrosine Kinases ; Receptor, Fibroblast Growth Factor, Type 3 ; Receptors, Fibroblast Growth Factor ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Translocation, Genetic