ABSTRACT
@#目的：通过向C57Bl/6J小鼠腹腔注射IFN-γ腺病毒（Ad-mIFN-γ）建立细胞因子释放综合征（CRS）的动物模型。方法：构建Ad-mIFN-γ及对照Ad-lacZ腺病毒载体，分别以MOI=100体外转染小鼠腹腔巨噬细胞，流式细胞术检测其对细胞mIFN-γ分泌的影响。将40只雌性C57Bl/6J小鼠按随机数字表法分为对照组、载体对照组、病毒低、中、高剂量组（每组8只），分别向各组小鼠腹腔注射PBS（200 μL）、Ad-lacZ（2×107 PFU/只）、Ad-mIFN-γ（5×106 PFU/只）、Ad-mIFN-γ（1.5×107 PFU/只）和Ad-mIFN-γ（2×107 PFU /只）。每日观测小鼠的体质量及生存情况；第3天时采用流式细胞术检测小鼠外周血和脾内单核细胞（CD11b+）、巨噬细胞（CD11b+/CD86+）比例，免疫荧光染色法检测脾内CD11b+的单核细胞比例；第9天时采用流式细胞术检测小鼠血清中细胞因子的分泌水平；第14天，采用颈椎脱臼法处死小鼠，H-E染色法观察小鼠肝、脾、肺和肾的病理和组织学变化。结果： Ad-mIFN-γ体外感染小鼠腹腔巨噬细胞，在第3天检测到巨噬细胞分泌mIFN-γ达到峰值（118.34±2.90）pg/mL，并在一周内持续高分泌mIFN-γ，Ad-lacZ对照组IFN-γ分泌水平较低后，第3天时为（0.17±0.08）pg/mL。小鼠腹腔注射Ad-mIFN-γ后，在14 d内病毒低、中剂量组无小鼠死亡，病毒高剂量组小鼠体质量持续减轻（P<0.001）；第3天，病毒高剂量组小鼠外周血和脾组织内单核细胞、巨噬细胞比例较对照组和中剂量组均显著增加（P<0.05或P<0.01）；第9天，病毒低、中、高剂量组小鼠血清中mIFN-γ、IL-6、单核细胞趋化蛋白-1（MCP-1）、IL-1、TNF-α等细胞因子的水平均显著升高（P<0.001）；10 d内病毒高剂量组小鼠死亡率达100%。组织病理检测可见病毒高剂量组小鼠的肝、脾、肺、肾组织有明显损伤。结论： Ad-mIFN-γ体外感染小鼠原代腹腔巨噬细胞后，可以快速分泌mIFN-γ；腹腔注射高剂量（2×107 PFU/只）Ad-mIFN-γ导致小鼠出现CRS典型表现，可作为CAR-T细胞治疗诱发CRS的动物模型。
ABSTRACT
The present study aimed to examine the value of ultrasonic soft markers in prenatal screening by analyzing the clinical outcome of fetuses with ultrasonic soft markers during the second trimester of pregnancy.A retrospective analysis was performed to evaluate the outcome of 591 fetuses with ultrasonic soft markers from January 2015 to August 2016 in Zhongnan Hospital of Wuhan University,China.It was found that 591 fetuses showed ultrasonic soft markers in 4927 cases with the occurrence rate being 12.0％.Among them,564 fetuses (95.4％) were delivered and the remaining 27 cases (4.6％) were aborted.Five hundred and sixty-seven cases had single ultrasonic soft marker,including echogenic intracardiac focus (n=343),mild renal pelvis dilatation (n=116),short long bones (n=72),single umbilical artery (n=31),mild lateral ventriculomegaly (n=21),choroid plexus cysts (n=19),and echogenic bowel (n=13),with the disappearing rates in pregnancy being 97.1％ (333/343),77.6％ (90/116),0％ (0/72),0％ (0/31),57.1％ (12/21),89.5％ (17/19) and 61.5％ (8/13) respectively.The rate of pregnancy termination due to single ultrasonic soft marker was 3.4％ (19/567),and that was 33.3％ (8/24) due to two ultrasonic soft markers with the difference being statistically significant (P＜0.05).The reasons of pregnancy termination included malformations (polycystic kidney,cleft lip and palate,congenital heart diseases,pcromphalus,hypospadias,hydrocephalus),chromosome abnormality,and stillbirth.It was concluded that single ultrasonic soft marker is usually transient manifestation in pregnancy.Without the other structural defects,single ultrasonic soft marker usually disappears spontaneously with favorable prognosis in a low-risk population.It is suggested that ultrasonic soft markers should be appropriately interpreted to avoid unnecessary invasive examination.
ABSTRACT
OBJECTIVE: One approach to identifying HIV-1 vaccine candidates is to dissect the natural antiviral immune response in treatment-naïve individuals infected for over ten years, considered slow progressor patients (SPs). It is suspected that SP plasma has strongly neutralizing antibodies (NAb) targeting specific HIV viral epitopes. METHODS: NAbs levels of 11 HIV-1-infected SPs were detected by PBMC-based neutralization assays. To investigate SP NAb epitope, this study used a biopanning approach to obtain mimotopes of HIV-1 that were recognized by SP plasma NAbs. IgG was purified from hightiter NAb SP plasma, and used as the ligand for three rounds of biopanning to select HIV-specific mimotopes from a phage-displayed random peptide library. Double-antibody sandwich ELISA, competitive inhibition assays, and peptide sequence analysis were used to evaluate the characteristics of phage-borne mimotopes. RESULTS: SPs had significantly more plasma neutralizing activity than typical progressors (TPs) (p = 0.04). P2 and P9 plasma, which have highest-titer HIV-NAb, were selected as ligands for biopanning. After three rounds of biopanning, 48 phage clones were obtained, of which 22 clones were consistent with requirement, binding with HIV-1 positive plasma and unbinding with HIV-1 negative plasma. Compared with linear HIV-1 protein sequence and HIV-1 protein structure files, only 12 clones were possible linear mimotopes of NAbs. In addition, the C40 clone located in gp41 CHR was found to be a neutralizing epitope, which could inhibit pooled HIV-1 positive plasma reaction. CONCLUSION: Biopanning of serum IgG can yield mimotopes of HIV-1-related antigen epitopes. This methodology provides a basis for exploration into HIV-1-related antigen-antibody interactions and furthers NAb immunotherapy and vaccine design.
Subject(s)
Humans , Antibodies, Neutralizing/immunology , Epitopes/immunology , HIV Antibodies/immunology , HIV Infections/immunology , HIV-1 , China , Cross Reactions , Enzyme-Linked Immunosorbent Assay , /immunology , /immunology , Peptide LibraryABSTRACT
The present study reports statistical medial optimization for chitinase production by a novel bacterial strain isolated from soil recently, which the name Chitinolyticbacter meiyuanensis SYBC-H1 is proposed. A sequential statistical methodology comprising of Plackett-Burman and response surface methodology (RSM) was applied to enhance the fermentative production of chitinase, in which inulin was firstly used as an effective carbon source. As a result, maximum chitinase activity of 5.17 U/mL was obtained in the optimized medium, which was 15.5-fold higher than that in the basal medium. The triplicate verification experiments were performed under the optimized nutrients levels which indicated that it well agreed with the predicted value.
Subject(s)
Carbon/analysis , Fermentation , Inulin/isolation & purification , Chitinases/analysis , Chitinases/isolation & purification , Data Interpretation, Statistical , Enzyme Activation , Methodology as a Subject , Process Optimization , MethodsABSTRACT
The study examined the inhibitory effect of Atractylodes macrocephala (AM) on the uterine contraction during premature delivery and explored its electrophysiological mechanism by studying the effects of AM on the Ca2+-activated K+ currents of pregnant human myometrial smooth muscle cells with or without the treatment with interleukin-6. Single cells were acutely isolated from pregnant human myometrial smooth muscles. Whole-cell Ca2+-activated K+ currents were recorded by using an Axopatchl-D amplifier. The cells were divided into three groups: group A in which AM was added into perfusate, group B, in which intefleukin-6 was added into perfusate) and group C in which AM was added into perfusate after addition of interleukin-6. IL-6 10 ng/mL inhibited Bkca by 36.9%±13.7% as compared with control (P<0.01). AM at 2 mg/mL raised Bkca by 36.7%±22.6% or 45.2%±13.7% with or without the treatment of IL-6, respectively (P<0.01). It is concluded that AM was able to enhance the Bkca of pregnant human myometrial smooth muscle cells treated or un- treated with interleukin-6 and its effect on the Bkca IL-treated cells was stronger that its effect on Bkca of untreated cells. Our results suggested that AM can help to maintain the membrane potentials and the resting status of pregnant human myometrial smooth muscle cells.
ABSTRACT
<p><b>OBJECTIVE</b>To identify and evaluate mutations in the RP1 gene among Chinese patients with retinitis pigmentosa (RP).</p><p><b>METHODS</b>Leukocyte DNA of 92 RP patients were collected in Hong Kong. Sequence changes of the entire coding region of the RP1 gene were examined using PCR, conformation sensitive gel electrophoresis and DNA sequencing.</p><p><b>RESULTS</b>In total, 1 nonsense mutation and 1 nonsense variant as well as 10 missense alterations were identified in the RP1 gene, among which, Arg677Ter was found in 1 RP patient and another nonsense variant, Arg1933Ter, was identified in 3 normal individuals and 1 patient with Stargardt's disease, suggesting its nonpathogenicity. Arg77Ter is expected to lead to large disruptions of the encoded protein.</p><p><b>CONCLUSIONS</b>The nonpathogenicity of Arg1933Ter indicates that the C-terminal 224 residues of RP1 protein may be not critical for RP1. The most C-terminal truncation previously reported was due to Tyr1053 (1-bp del) and occurred in RP patients. Thus RP can be caused by reduction in the level of the region of RP1 protein after codon 1052 but before 1933. To ascertain such a proposition, genotypes of more RP patients may reveal more RP causative mutations and more sequence alterations different than those of other ethnic groups.</p>