The effects of Guizhi Fuling capsule and its active ingredient combination within different concentration on SPL proliferate were observed by MTT method. The ratio of CD80/86, CD3CD25 and CD3CD69 was used to evaluate cell activation effects of Guizhi Fuling capsule and its active ingredient combination by FCM. Guizhi Fuling capsule with concentration of 400 mg · L(-1)can promote spleen lymphocyte proliferation, as well as the active ingredient combination, which showed the obvious dose-effect relationship. Compared with control group, the difference has statistical significance (P≤0.01). The result of FCM showed that Guizhi Fuling capsule and its active ingredient combination can promote CD80 and CD86 expression on spleen lymphocyte, and also can increase CD25 and CD69 ratio between spleen CD3+ cells. Compared with control group, the difference has statistical significance (P≤0.01). Thus, Guizhi Fuling capsule and its active ingredient combination may have immune-modulate effects, and the mechanism may have a close relationship with the lymphocyte activation.
Animals ; Capsules ; Drugs, Chinese Herbal ; analysis ; pharmacology ; Immunologic Factors ; pharmacology ; Lymphocyte Activation ; drug effects ; Male ; Mice ; Mice, Inbred BALB C ; T-Lymphocytes ; drug effects ; immunology

To study effect of vitrification state of protective solutions on recovery of red blood cells after lyophilization, four protective solutions composed of isotonic buffers containing 7% DMSO (v/v) and 20%, 30%, 40% or 50% polyvinylpyrrolidone (PVP) (w/v) were adopted. Vitrification state of protective solutions was examined first when white ice crystal appeared in any protective solution during freezing or thawing, if the used solution was not a vitrification solution. Red blood cells were lyophilized in MINILYO45 freeze-dryer after washing, mixing with protective solutions and prefreezing. After lyophilization, the samples were quickly rehydrated by 37 degrees C rehydration solution. The results showed that in vitrification and devitrification experiments, white ice crystal appeared in solution of 20% PVP + 7% DMSO and 30% PVP + 7% DMSO during freezing and thawing; vitrification appeared in solution of 40% PVP + 7% DMSO during freezing, but devitrification appeared during thawing; vitrification appeared in solution of 50% PVP + 7% DMSO during freezing and thawing. After rehydration, the recoveries of red blood cells and hemoglobin in 40% PVP + 7% DMSO group were (81.36 +/- 14.94)% and (77.54 +/- 12.86)%, which were significantly higher than that in 20% PVP + 7% DMSO, 30% PVP + 7% DMSO and 50% PVP + 7% DMSO groups (P < 0.01). The concentration of free hemoglobin in 40% PVP + 7% DMSO group was also significantly lower than that in other three groups (P < 0.01). With increase of PVP concentration in protective solutions, vitrification state and protective effect of these solutions also increased; when concentration of PVP in protective solution was 40% though it was not a vitrification solution, the effect of lyophilization was the best; but when concentration of PVP further increased to 50%, though it was a vitrification solution, the effect decreased. It is concluded that excessive vitrification state could not benefit lyophilization of red blood cells.
Cryoprotective Agents ; pharmacology ; Dimethyl Sulfoxide ; pharmacology ; Dose-Response Relationship, Drug ; Erythrocytes ; cytology ; drug effects ; ultrastructure ; Freeze Drying ; methods ; Humans ; Microscopy, Electron ; Povidone ; pharmacology

Ductus Arteriosus ; abnormalities ; Fatal Outcome ; Female ; Heart Septal Defects, Atrial ; etiology ; Humans ; Infant, Newborn ; Rubella Syndrome, Congenital ; complications

OBJECTIVETo probe into a better therapy for primary trigeminal neuralgia.

METHODSEighty-six cases were randomly divided into an observation group (n = 46) and a control group (n = 40). The observation group were treated with the three-combination needling method, i. e. acupuncture, acupoint-injection and fire-needle therapy, and the control group with acupuncture and acupoint-injection. After treatment of 2 courses, their therapeutic effects were assessed.

RESULTSThe total effective rate of 93.5% and the cured rate of 60.9% in the observation group were better than 65.0% and 22.5% in the control group, with significant differences (both P < 0.01).

CONCLUSIONThe three-combination needling method has obvious clinical therapeutic effect on primary trigeminal neuralgia.

Acupuncture Therapy ; methods ; Adolescent ; Adult ; Female ; Humans ; Male ; Middle Aged ; Trigeminal Neuralgia ; diagnosis ; therapy

OBJECTIVETo prepare the genetic engineering regulatory T cells (Treg) via the self-inactivating (SIN) lentiviral vectors carrying Foxp3 gene, and assay the phenotype and abilities of its proliferation and immunosuppression.

METHODSThe bicistronic SIN lentiviral transfer plasmid containing Foxp3 gene and internal ribosomal entry site-green fluorescent protein gene (IRES-GFP) was constructed. Human embryonic kidney 293T cells were co-transfected using liposome by lentiviral packing system, which included the packaging plasmid Delta NRF, the transfer plasmid and the envelope plasmid VSVG. The efficiency of gene transduction and the expressions of Foxp3, CD25, GITR, CTLA-4 of CD4(+)CD25(-) T cells, which were isolated by magnetic beads from the spleen, and then co-cultured with 293T cells, were detected by flow cytometry (FCM). The proliferative and suppressive capacities of transduced T cells were estimated by Cell Count Kit-8 (CCK-8) and the cytokine production was performed by ELISA.

RESULTSThe lentiviral transfer plasmid pXZ208-Foxp3-IRES-GFP was successfully constructed, the virus titers were above 10(6) IU/ml in the supernatant. pXZ208-IRES-GFP was used as control group. After cocultured, the CD4(+)CD25(-) T cells expressed significantly higher Foxp3, CD25, GITR and CTLA-4 in experimental group than in control group. Upon stimulation with anti-CD3 epsilon and APCs, the proliferative capacity of Foxp3-transduced T cells and the production of IL-2, IL-4, IL-10, IFN-gamma were significantly lower than those in control group (P < 0.01); Foxp3-transduced T cells also significantly inhibited the proliferation of CD4(+)CD25(-) T cells.

CONCLUSIONSThe genetic engineering Treg mediated by SIN lentiviral vectors are successfully constructed and their phenotype and function are similar to natural CD4(+)CD25(+) Treg.

Animals ; Cell Proliferation ; Cells, Cultured ; Forkhead Transcription Factors ; genetics ; metabolism ; Genetic Engineering ; Genetic Vectors ; HEK293 Cells ; Humans ; Lentivirus ; genetics ; Mice ; Mice, Inbred BALB C ; Phenotype ; T-Lymphocytes, Regulatory ; cytology ; immunology ; metabolism ; Transfection

OBJECTIVETo study the plasma protein binding rate of methyl protodioscin.

METHODThe ultrafiltration was employed to determine the plasma protein binding rate of methyl protodioscin. The plasma concentrations of methyl protodioscin were measured by HPLC-MS-MS.

RESULTThe plasma protein binding rate of methyl protodioscin with rat plasma at the concentration of 20.0, 100 and 200 microg x mL(-1) were (94.6 +/- 0.16)%, (91.6 +/- 0.35)% and (86.10 +/- 0.60)%, respectively, while the plasma protein binding rate of methyl protodioscin with normal human plasma at the above concentrations were (82.11 +/- 5.12)%, (84.54 +/- 0.32)% and (88.52 +/- 1.02)%, respectively.

CONCLUSIONThe binding rate of methyl protodioscin with plasma protein is high.

Animals ; Antineoplastic Agents ; metabolism ; Blood Proteins ; metabolism ; Calibration ; Chromatography, High Pressure Liquid ; Diosgenin ; analogs & derivatives ; metabolism ; Female ; Humans ; Male ; Protein Binding ; Rats ; Saponins ; metabolism ; Sensitivity and Specificity ; Tandem Mass Spectrometry ; Ultrafiltration

Objective To investigate clinical symptoms,pathophysiology and brain imaging features of Chinese familial cerebral cavernous angiomas.Methods Head MRI examination,clinical and pathophysiological examination were performed in a Chinese family with one proband of cerebral cavernous malformation.The disease atlas of the family was drawn.The patients indicative of a surgery underwent resection of hemangioma whose pathophysiology and microstructure were observed.Results Nine familial cerebral cavernous angiomas patients were found to have multiple intracranial lesion in the 18 family members,the penetrance being 50%,conforming to the feature of autosomal incomplete dominance inheritance.Four patients with skin cavernous hemangioma had familial cerebral cavernous angiomas.MRI was the most sensitive modality for the diagnosis of cavernous angioma.With T_2-Weighted sequences,the lesion was typically characterized by an area of mixed signal intensity,with a central reticulated core and a peripheral rim of decreased signal intensity related to deposition of hemosiderin.Gradient-echo(GRE)MRI could find microcavernous hemangiomas that would not be found in other sequences.Cavernous angiomas were typically discrete multilobulated berrylike lesions that contained hemorrhage in various stages of evolution.Histological homogeneity and overlap with other vascular malformations such as capillary telangictasia was common.Cavernous angiomas were composed of endothelial-linked sinusoidal spaces not separated by significant amounts of neural tissue.Hemorrhagic residua were common.Clots at different stages of evolution within the lesion were seen.The basic membranes of sinus became thick and soft.Parts of it were layered.Conclusions Familial cerebral cavernous angiomas is an autosomal incomplete dominance inheritance disease.Cavernous angiomas are composed of endothelial-linked sinusoidal spaces not separated by significant amounts of neural tissue.There are more than one focus in every patients and the skin cavernous angiomas is the foundation of diagnosing familial cerebral cavernous angiomas.Gradient-echo imagine sequence MRI(3.0 T)could be the"golden standard".

To study effect of pre-freezing temperature and lyophilizer shelf temperature on recovery of human red blood cells after lyophilization and determine solidifying temperature of this lyophilization system, the protective solution composed of 7% DMSO, 40% polyvinylpyrrolidone (PVP) and isotonic buffer were adopted to lyophilize red blood cells at different pre-freezing temperatures or shelf temperatures. At first, fresh whole blood was centrifugated, washed and equilibrized to prepare concentrated red blood cells. Then concentrated red blood cells were mixed with the protective solution at 1:3 and pre-freezed at different temperature (-20, -35, -45, -80 or -196 degrees C) before lyophilization in lyophilizer. To study effect of shelf temperature on lyophilization of red blood cells, red blood cells were lyophilized at different shelf temperature after pre-freeze at -80 degrees C. After lyophilization, the samples were quickly rehydrated by 37 degrees C rehydration solution. The results showed the recovery rate of red blood cells and hemoglobin after pre-freeze at different temperature and lyophilization were > 85% and > 75%, there was not significant difference among these groups, but the concentration of free hemoglobin in -196 degrees C group was significantly higher than that in other groups (P < 0.01). With decreasing of shelf temperature, the lyophilizing time was also prolonged. When shelf temperature was > or = -25 degrees C, samples were not fully lyophilized; when shelf temperature was < or = -30 degrees C, the recovery rate of red blood cells and hemoglobin after lyophilization and rehydration were above 90%; after washed to isotonic state, the recovery rate of hemoglobin of the four groups was similar to each other. In conclusion, only when pre-freezing temperature is between -20 and -80 degrees C and the lyophilizer shelf temperature is < or = -30 degrees C, the effect of lyophilization is better, but the effect of excessively low pre-freezing temperature may even be worse.
Blood Preservation ; Erythrocytes ; cytology ; Freeze Drying ; Hemoglobins ; analysis ; Humans ; Temperature

OBJECTIVETo explore the influence of the lentiviral vector mediated mouse genetic engineering regulatory T cells (Treg) infusion on post-allogeneic bone marrow transplantation (allo-BMT) acute graft-versus-host disease (GVHD) in mice.

METHODSLentivirus-mediated Forkhead box P3 (Foxp3) gene was transduced into BALB/c mice CD4(+)CD25(-) T cells (Treg) to construct engineered Tregs in vitro. An allo-BMT model of BALB/c to C57BL/6 mice was established. After irradiation, the recipients were injected with donor cells plus the genetic engineering Tregs. Survival time, clinical GVHD score, histopathological findings, activation of donor T cells or serum levels of inflammatory cytokines were observed after allo-BMT.

RESULTSThe mean survival times for radiation alone group (Gp I), transplantation control group (Gp II), engineering Treg infusion group (Gp III) and empty vector control group (Gp IV) were (8.8 ± 0.6) d, (36.7 ± 2.5) d, (51.6 ± 4.0) d and (34.1 ± 2.3) d, respectively. The survival time was significantly longer in Gp III than in other groups (P < 0.05). Histopathological finding in several target organs (skin, liver and small intestine) confirmed the presence of severe GVHD in Gp II and Gp IV, while no histological signs of GVHD were observed in long survival recipients in Gp III, and clinical GVHD scores in Gp III were significantly lower than that in Gps II and IV. The numbers of donor T cells and the percentage of IFN-producing donor T cells in the spleen of recipients in Gp III were significant lower than those in Gps II and IV at days 3 and 4, and at day 3 after transplantation, respectively (P < 0.05). The serum levels of IFN-γ, IL-2 and TNF-α were increased at day 21 to 28 after transplantation in all groups. The peak concentrations of IFN-γ, IL-2 and TNF-α in Gp III were significantly lower than those in Gps II and IV control groups at day 21 (P < 0.05).

CONCLUSIONCo-injection of genetic engineering Treg can efficiently prevent recipients from lethal GVHD after allo-BMT in mice by inhibiting the early activation and expansion of donor T cells and reducing the serum levels of inflammatory cytokines.

Animals ; Bone Marrow Transplantation ; adverse effects ; Cytokines ; blood ; Female ; Forkhead Transcription Factors ; genetics ; Genetic Engineering ; Genetic Vectors ; Graft vs Host Disease ; immunology ; Lentivirus ; Lymphocyte Activation ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; T-Lymphocytes, Regulatory ; cytology ; immunology ; Transduction, Genetic ; Transplantation, Homologous

The aim of this study was to investigate the expression of Th17 cells and regulatory T (Treg) cells in peripheral blood of patients with immune thrombocytopenia (ITP) and to clarify the role of the Th17/Treg cell ratio imbalance in pathogenesis of ITP. Patients were divided into the pre-treatment group (active group) (n = 38) and post-treatment group (remission group) according to the platelet count and curative effect. Post-treatment group was further divided into remission group (n = 24), partial remission group (n = 10), and non-remission group (n = 4). 30 healthy subjects were enrolled in control group. Flow cytometry was used to detect the percentages of peripheral blood Th17 cells and Treg cells in CD4(+) T cells from ITP patients and controls respectively. The results showed that the percentages of Th17 cells in active group and non-remission group were significantly higher than those in control group (p < 0.05). The percentages of Th17 cells in remission group, partial-remission group were also higher than those in control group, but there were no statistically significant differences between these groups. The percentage of Th17 cells in remission group was lower than that in active group, but there was also no statistically difference between two groups. The percentages of Treg cells in active group, partial-remission group and non-remission group significantly decreased, compared with in control group (p < 0.01). The percentage of Treg cells in remission group was lower than that in control group, but there was no statistically significant difference. The ratio of peripheral blood Th17/Treg cells in active group, partial-remission group and non-remission group was higher, as compared with in control group. The ratio of peripheral blood Th17/Treg cells in remission group was higher than that in control group, but there was no statistically difference between two groups. It is concluded the percentage of Th17 cells and the ratio of Th17/Treg cells are higher in active group. The percentage of Treg cells is low in active group, partial remission and non-remission groups. The imbalance of Th17/Treg ratio may play a critical role in ITP pathogenesis.
Adolescent ; Adult ; Aged ; Blood Cell Count ; Case-Control Studies ; Female ; Flow Cytometry ; Humans ; Male ; Middle Aged ; Purpura, Thrombocytopenic ; blood ; T-Lymphocytes, Regulatory ; immunology ; Th17 Cells ; immunology ; Young Adult