OBJECTIVEThis study investigates the immune status of T helper (Th) 17 cells in mice with periodontitis.

METHODSSeven-week-old C57BL/6 female mice were randomly divided into the control and periodontitis groups. Each group comprisedfour mice. Experimental periodontitis was induced in mice by oral infection with Porphyromonas gingivalis in the periodontitis group. Phosphate-buffered saline solution was used in the control group. All mice were sacrificed 4 weeks after the last P. gingivalis infection. CD4⁺retinoid-related orphan receptor (ROR) γτ⁺(Th17) cells were analyzed by flow cytometry. The protein expression of Th17 cell-related cytokine interleukin (IL)-17A was detected by enzyme-linked immunosorbentassay (ELISA).

RESULTSFlow cytometry showed that the percentage of CD4⁺RORγτ⁺(Thl7) cells in all CD4⁺ cells and the cell number in gingival tissues, cervical lymph nodes (CLNs), and the peripheral blood obviously increased (P < 0.01) in the periodontitis group. ELISA showed that compared with the control group, the periodontitis group exhibited an obvious increase in the protein expression of IL-17A (P < 0.05).

CONCLUSIONTh17 cell-mediated cell response is enhanced, and the gingival tissues, CLNs, and the peripheral blood are probably the main locations of Thl7 cell-mediated cell response during the development of periodontitis.

Alveolar Bone Loss ; Animals ; Cytokines ; Female ; Flow Cytometry ; Gingiva ; Interleukin-17 ; metabolism ; Mice ; Mice, Inbred C57BL ; Periodontitis ; metabolism ; Porphyromonas gingivalis ; Random Allocation ; T-Lymphocytes, Helper-Inducer

Objective:To investigate regulatory T(Treg)cell-mediated immune response in mice with periodontitis.Methods:8 sev-en-week old C57BL/6 female mice were randomly divided into 2 groups(n =4).Periodontitis was induced in mice by oral infection with Porphyromonas gingivalis(P.gingivalis).Carboxymethyl cellulose(CMC)was used for the sham control mice.All mice were sacrificed 4 weeks after the last P.gingivalis infection.CD4 +forkhead box P3 (Foxp3)+ cells were analyzed by flow cytometry.The protein ex-pression of TGF-1 and IL-1 0 was detected by ELISA.Results:Periodontitis mice showed an obvious decrease in the percentage of CD4 +Foxp3 + cells in CD4 + T cells in gingival tissues,cervical lymph nodes(CLNs)and peripheral blood(P <0.01 ).Moreover,peri-odontitis mice had an obvious increase in the expression of TGF-1 and IL-1 0 in gingival tissues,CLNs and serum(P <0.05).Conclu-sion:Treg cell-mediated immune response is weakened in the mice with periodontitis.Gingival tissues,CLNs and the peripheral blood are probably the main source of Treg cells during periodontitis.

OBJECTIVETo prepare colon target pellets of Pulsatilla total saponins.

METHODPulsatilla total saponins-hydroxypropyl-beta-cyclodextrin inclusion was prepared by the water solution-mixing method. Then plain pills of inclusion were prepared by the granulation-spheronization method, and coated by Glatt fluid bed.

RESULTThe dissolution of plain pills of Pulsatilla total saponins at 2 h was 16.0%, while that of plain pills of inclusion at 0.5 h was 91.9%. With Eudragit S100 as the coating material, TEC as the plasticizer and talcum power as the anti-adherent, when the coating weight was 12%, the coating efficiency was high, with almost no bonding and drug release of coated pellets in artificial gastric juice for 2 h. The accumulated drug release in artificial intestinal fluid for 4 h was less than 15%, and that in artificial colon fluid for 4 h was more than 90%.

CONCLUSIONCoated pellets of Pulsatilla total saponins-hydroxypropyl-beta-cyclodextrin inclusion showed a good colon targeted drug release in vitro, thus could be further developed to be oral colon targeted preparations.

2-Hydroxypropyl-beta-cyclodextrin ; Absorption ; Biomimetic Materials ; metabolism ; Colon ; metabolism ; Drug Compounding ; methods ; Drug Implants ; Gastric Juice ; metabolism ; Humans ; Pulsatilla ; chemistry ; Saponins ; chemistry ; metabolism ; Surface Properties ; beta-Cyclodextrins ; chemistry

OBJECTIVETo investigate the magnetic resonance imaging (MRI) feature of multiple cerebral sclerosis (MS) for better understanding and diagnosis of this disease.

METHODSThe MRI data of 32 patients with MS were reviewed. Conventional scanning with T1WI, T2WI, Flair sequence was performed, and 26 patients underwent Gd-DTPA enhanced scanning. The MS plaques were analyzed for their locations, sizes, shapes, MR signals and enhanced features, space-occupying signs, and the related corpus callosum changes and brain atrophy. Descriptive statistical method was used for all the data.

RESULTSMRI identified MS lesions in the brain in 30 cases, with the sensitivity of 93.75%. All the MS patients had multiple lesions with predilection sites of the cortical/juxtacortical and periventricle areas, the centrum semiovale, and the corpus callosum. Most of the MS plaques were round or oval of different sizes. Bilateral lesions were almost symmetrical in distribution. Twenty patients had "rectangular demyelination" and 12 had "dirty white matter" signs, and 11 had both manifestations. The lesions were isointense, slightly hypointense or hypointense on T1WI, and hyperintense on T2WI and Flair sequences. Most of the MS plaques presented no enhancement, with occasional nodular or circular enhancement. No or slight space-occupying effect was found in the plaques. Of the 28 MS patients undergoing sagittal scanning of the corpus callosum, 17 presented with abnormal signals, with the sensitivity of 60.71% (17/28). Five patients had corpus callosum atrophy, and 10 had brain atrophy of different degrees.

CONCLUSIONThese results suggest that the corpus callosum is often compromised by the MS lesions to present diffusive, nodular, radiating signal abnormalities and irregular ependymal thickening, which can be most obvious with sagittal FLAIR imaging.

Adolescent ; Adult ; Brain ; pathology ; Corpus Callosum ; pathology ; Female ; Humans ; Magnetic Resonance Imaging ; methods ; Male ; Middle Aged ; Multiple Sclerosis ; diagnosis ; Retrospective Studies ; Young Adult

Diagnosis, Differential ; Female ; Humans ; Laparotomy ; methods ; Pregnancy ; Pregnancy, Ectopic ; diagnosis ; surgery ; Retroperitoneal Space ; diagnostic imaging ; Tomography, X-Ray Computed ; Ultrasonography

Objective To investigate the click and tone burst evoked auditory brainstem responses (ABR)in normal Wistar rat,and to establish the standards of ABR testing method,and to provide a reference for studies rat audition.Methods Fifteen male Wistar rats(30 ears)were used in this sutdy.The latency and amplitude of ABR e-voked by click and TB at 80,50 and 20 dB SPL were measured.Results The occurrence rate of wave Ⅱand Ⅳat low levels(20 dB SPL)was nearly the same according to the amplitude.The cABR (dB peSPL)threshold was 21.83± 4.45 and tbABR (dB SPL)thresholds were 2.02±0.09,2.88±0.16,3.77±0.25,4.69±0.29,and 5.78±0.41, respectively.80 dB stimulus evoked cABR (peSPL)wave I,I b,II,III,IV and V latency (ms)were 1.76±0.12, 2.13±0.11,2.67±0.16,3.49±0.28,4.39±0.29,and 5.45±0.41,respectively.tbABR (SPL)of wave I,Ib, II,III,IV and V latency (ms)at 4 kHz were 2.02±0.09,2.88±0.16,3.77±0.25,4.69±0.29,and 5.78± 0.41,respectively.At 8 kHz they were 1.76±0.07,2.28±0.10,2.63±0.16,3.49±0.21,4.44±0.28,and 5.48±0.43;while at 12 kHz were1.76±0.08,2.24±0.12,2.61±0.25,3.53±0.25,4.46±0.32,and 5.52± 0.45;at 16 kHz were 1.79±0.10,2.25±0.12,2.70±0.18,3.62±0.27,4.52±0.37,and 5.61±0.49;at 24 kHz were 1.75±0.09,2.27±0.11,2.67±0.16,3.60±0.27,4.52±0.38,and 5.60±0.51;at 32 kHz were 1.77±0.10,2.24±0.12,2.64±0.20,3.59±0.34,4.52±0.40,and 5.61±0.52,respectively.Conclusion Wave Ⅳ was the best wave to determine threshold of click and tone burst evoked auditory brainstem response in rat.

De novo variants (DNVs) are one of the most significant contributors to severe earlyonset genetic disorders such as autism spectrum disorder, intellectual disability, and other developmental and neuropsychiatric (DNP) disorders. Presently, a plethora of DNVs have been identified using next-generation sequencing, and many efforts have been made to understand their impact at the gene level. However, there has been little exploration of the effects at the isoform level. The brain contains a high level of alternative splicing and regulation, and exhibits a more divergent splicing program than other tissues. Therefore, it is crucial to explore variants at the transcriptional regulation level to better interpret the mechanisms underlying DNP disorders. To facilitate a better usage and improve the isoform-level interpretation of variants, we developed NeuroPsychiatric Mutation Knowledge Base (PsyMuKB). It contains a comprehensive, carefully curated list of DNVs with transcriptional and translational annotations to enable identification of isoformspecific mutations. PsyMuKB allows a flexible search of genes or variants and provides both table-based descriptions and associated visualizations, such as expression, transcript genomic structures, protein interactions, and the mutation sites mapped on the protein structures. It also provides an easy-to-use web interface, allowing users to rapidly visualize the locations and characteristics of mutations and the expression patterns of the impacted genes and isoforms. PsyMuKB thus constitutes a valuable resource for identifying tissue-specific DNVs for further functional studies of related disorders. PsyMuKB is freely accessible at http://psymukb.net.

OBJECTIVETo compare the effects of different types of feeder cells on supporting undifferentiation and high proliferation of human embryonic stem cells (hESC).

METHODShESC were seeded on mouse embryonic fibroblasts (MEF), human marrow stromal cells (hMSC), and human foreskin fibroblasts (hFF), respectively. Colony number, cell quantity after digestion, and survival rate were observed by alkaline phosphatase (AP) staining and Trypan blue, and the biological properties of hESC after 5 passages were observed by immunofluorescence staining.

RESULTSAlthough all the three feeder layers could support the formation of hESC colonies and maintain pluripotency, the morphology of colonies on different feeder layers remarkably varied. The stage-specific embryonic antigen-3 and AP staining were positive on three types of feeders. The number of colonies, number of cells produced, and cell survival rates were significantly higher on MEF than on human feeder cells (P < 0.01). Furthermore, the number of AP-positive colonies and cell quantity were also significantly higher on hMSC than on hFF (P < 0.01).

CONCLUSIONSAll three types of feeder cells are able to support the growth of hMSC, although MEF are more favourable for the proliferation. Two types of human feeder cells lay the foundation for the removal of animal-derived hESC culture system. hMSC is superior to hFF in supporting the proliferation of hESC.

Animals ; Antigens, Tumor-Associated, Carbohydrate ; metabolism ; Coculture Techniques ; methods ; Embryonic Stem Cells ; Feeder Cells ; Fibroblasts ; Humans ; Mice ; Stage-Specific Embryonic Antigens ; metabolism

The study was designed to investigate annexin II resulting in molecular pathological mechanism of the primary fibrinolysis and establish annexin II vector model for further research on disturbance of coagulation. A target gene was amplified from human umbilical vein endothelial cells (HUVEC) by RT-PCR. Annexin II gene fragment was purified and ligated with molecular biological recombinant technology. The recombinant of plasmid annexin II was transfected into HL-60 cells and its distribution in the cell and structure characteristics of annexin II protein were evaluated by multi-photon excitation laser scanning microscope. By means of flow cytometry (FCM) and Werstern blot technique, the protein expression was qualitatively and quantitatively analyzed. Transfected cells were treated in vitro with annexin II antisense oligonucleotide (AS) targeting to the start site of annexin II cDNA. The results showed that the recombinant pZeoSV2(+)/ANN II was constructed successfully and expressed in HL-60 cells. The protein expression was distributed on the surface of cell by fluorescence assay. After transfection for 48 hours, the cells occurred higher level of expression. The level of the plasmin was significantly enhanced in the present of annexin II. The FCM and Western blot analysis showed the annexin II expression was similar both in transiently and stably transfected in HL-60 cells. Annexin II antisense oligonucletide and McAb significantly inhibited the activity of plasminogen. It was concluded that annexin II plays an important role in the fibrinotysis. Annexin II vector was defined as a expression tool for further studying fibrinolysis and coagulopathy in malignant disease.
Annexin A2 ; genetics ; physiology ; Endothelium, Vascular ; chemistry ; cytology ; Fibrinolysis ; Genetic Vectors ; HL-60 Cells ; Humans ; Oligonucleotides, Antisense ; pharmacology ; Polymerase Chain Reaction ; Recombinant Proteins ; biosynthesis

OBJECTIVETo investigate the effect and its mechanism of reducing graft-versus-host disease (GVHD) by in vitro blockade of CD(40)-CD(40)L pathway in vitro, the donor T lymphocytes cultured in vitro with anti-CD(40)L mAb were transfused in bone marrow transplantation (BMT) GVHD mouse model.

METHODSC57BL/6(H-2b) spleen T cells were isolated as responder cells, and BALB/c(H-2d) spleen cells as stimulator cells. They were cocultured with or without Anti-CD(40)L mAb as anti-CD(40)L mAb group and control group, respectively. At day 5, the mixed lymphocyte response (MLR)-culture cells mixed with bone marrow cells and transfused respectively into the TBI conditioned recipient mice. The mice were divided into two groups: group A, bone marrow cells (2 x 10(6)) and spleen T lymphocytes (2 x 10(6)) from MLR control group; group B, bone marrow cells (2 x 10(6)) and spleen T lymphocytes (2 x 10(6)) from MLR anti-CD(40)L mAb group. The GVHD incidence and hematopoietic reconstitution were observed. Peripheral blood sera and spleen cells of the recipients mice were harvested at scheduled time points for the measurement of cytokines and T cell immunophenotyping with flow cytometry.

RESULTSThe incidence of GVHD in group A was 100% (10/10), and in group B was 20% (2/10). The percentage of H-2D(b) positive cells in group B (n = 8) was (93.54 +/- 2.32)% at day 40 after transplantation. The levels of cytokines in serum from group B were significantly lower than those from group A (P < 0.05). The expressions of CD(4)(+), CD(8)(+), CD(4)(+)CD(25)(+), CD(8)(+)CD(25)(+), CD(4)(+)CD(69)(+), CD(8)(+)CD(69)(+) and CD(4)(+)CD(40)L(+) were lower in group B than in group A (P < 0.05). The expressions of CD(8)(+)CD(40)L(+) and CD(4)(+)CD(45)RA(+) were similar in the two groups (P > 0.05).

CONCLUSIONBlockade of CD(40)-CD(40)L interaction in vitro could induce immune tolerance in vivo, reduce aGVHD in aGVHD mice model and form chimerism, which was mediated by inhibiting the Th1 and Th2 cytokines production, inducing tolerance of CD(4)(+) and CD(8)(+) cells to alloantigens. The obstruction of T cells activation after tolerance happened mainly at the early and mature phase of T cells activation. These provided the experimental basis for the use of anti-CD(40)L mAb in the clinical transplantation to prevent aGVHD.

Animals ; Antibodies, Monoclonal ; therapeutic use ; Bone Marrow Transplantation ; adverse effects ; CD40 Antigens ; physiology ; CD40 Ligand ; immunology ; physiology ; Graft vs Host Disease ; prevention & control ; Interleukin-10 ; blood ; Interleukin-4 ; blood ; Male ; Mice ; Mice, Inbred C57BL