BACKGROUND: We developed a single-color multitarget flow cytometry (SM-FC) assay, a single-tube assay with graded mean fluorescence intensities (MFIs). We evaluated the repeatability of SM-FC, and its correlation with multicolor flow cytometry (MFC), to assess its application as a routine FC assay. METHODS: We selected CD19, CD3, CD4, and CD8 as antigen targets to analyze a lymphocyte subset. MFIs were graded by adjusting monoclonal antibody (mAb) volumes to detect several cell populations. Dimly labeled mAb was prepared by decreasing mAb volume and the optimum diluted volume was determined by serial dilution. SM-FC repeatability was analyzed 10 times in 2 normal controls. The correlation between SM-FC and MFC was evaluated in 20 normal and 23 patient samples. RESULTS: CV values (0.8-5.0% and 1.3-4.1% in samples 1 and 2, respectively) acquired by SM-FC with CD3-fluorescein alpha-isothyocyanate (FITC)dim+CD4-FITCbright and with CD19-FITCdim+CD3-FITCbright showed good repeatability, comparable to that acquired by MFC (1.6-3.7% and 1.0-4.8% in samples 1 and 2, respectively). Excellent correlation was observed between the 2 methods in the 20 normal samples (B cells, T cells, non-Thelper cells, and Thelper cells; r2=0.87, 0.97, 0.97, and 0.98, respectively; P<0.05). There were also linear relationships between SM-FC with CD19-FITCdim+CD3-FITCbright and CD8-PEdim+CD4-PEbright, and MFC, in the 23 patient samples (B cells, T cells, Tcytotoxic cells, and Thelper cells; r2> or =0.98, 0.99, 0.99, and 0.99, respectively; P<0.05). CONCLUSIONS: The multicolor, single-tube SM-FC technique is a potential alternative tool for identifying a lymphocyte subset.
Antibodies, Monoclonal/chemistry/*immunology ; Antigens, CD19/chemistry/metabolism ; Antigens, CD3/chemistry/metabolism ; Antigens, CD4/chemistry/metabolism ; Antigens, CD8/chemistry/metabolism ; B-Lymphocyte Subsets/immunology/metabolism ; Color ; Flow Cytometry/*methods ; Fluorescein-5-isothiocyanate/*chemistry ; Humans ; T-Lymphocyte Subsets/immunology/metabolism

The pathogenesis of some autoimmune diseases has been considered to be related to abnormal differentiation of T cell subsets. This study was aimed at investigating the change of Th1-like and Th2-like cells balance in ITP children, and analyzing the role of T cell subsets disequilibrium in the pathogenesis of ITP. Peripheral blood T cells were collected from 30 ITP patients, the T-cells were isolated and purified. The ratios of Th/Tc, Th1/Th2 and Tc1/Tc2 in peripheral blood T cells were analyzed by immunofluorescence staining and bicolor flow cytometry (FCM) in vitro. The results showed that as compared with the ratios of Th1/Th2 (48.76% +/- 6.17%) and Tc1/Tc2 (18.90% +/- 4.12%) in healthy children, the ratios of Th1/Th2 (56.21% +/- 5.95%) and Tc1/Tc2 (23.09% +/- 3.31%) in ITP children increased obviously. FCM analysis revealed that average percentages of Th, Th1, Th2, Tc, Tc1 and Tc2 were 22.31% +/- 6.51%, 21.92% +/- 6.42%, 0.39% +/- 0.14%, 31.12% +/- 6.15%, 30.95% +/- 5.45% and 1.34% +/- 0.84% in ITP children versus 39.24% +/- 5.82%, 39.01% +/- 5.47%, 0.80% +/- 0.16%, 30.25% +/- 5.63%, 28.72% +/- 5.20% and 1.52% +/- 0.68% in healthy children. The average percentages of Th, Th1 and Th2 decreased obviously, while the average percentages of Tc, Tc1 and Tc2 did not change. It is concluded that the ratios of Th1/Th2 and Tc1/Tc2 in peripheral blood T cells increase obviously in ITP children and the cellular immunity in ITP children shifts to Th1 type immunity superiority, which suggest that the abnormal differentiation of T cell subsets may play an important role in the pathologic process of ITP.
Child ; Child, Preschool ; Female ; Humans ; Infant ; Male ; Purpura, Thrombocytopenic, Idiopathic ; immunology ; T-Lymphocyte Subsets ; immunology ; metabolism ; pathology ; T-Lymphocytes, Cytotoxic ; chemistry ; Th1 Cells ; immunology ; Th2 Cells ; immunology

OBJECTIVETo observe effect of oral scorpio and scolopendra powder on T-cell subsets in peripheral blood and intestine from rats with collagen induced arthritis (CIA).

METHOD60 rats were randomly divided into 6 groups: normal control group, model control group, low-dose scorpio and scolopendra group, middle-dose scorpio and scolopendra group, high-dose scorpio and scolopendra group, and type II collagen group. Rat's rheumatoid arthritis was induced by collagen II (C II). Level of T-cell subsets from peripheral blood and intestine was measured by flow cytometry.

RESULTCD4+ T cellular level was obviously increased (P < 0.05 or P < 0.01) or kept increased tendency in peripheral blood and intestine from the model group compared with that of the normal group, while the ratio of CD4+/CD8+ in intestine was obviously descent but the contrary in peripheral blood (P < 0.05 or P < 0.01). CD4+, CD8+ T cellular level in intestine were obviously descent and the ratio of CD4+ /CD8+ increased in all treated groups when compared with in the model group (P < 0.05 or P < 0.01). However, CD4+ T cellular level and the ratio of CD4+/CD8+ in peripheral blood were remarkablely decreased.

CONCLUSIONThe mechanism that scorpio and scolopendra could treat rat's rheumatoid arthritis may be regulating balance of T-lymphocyte subsets in peripheral blood and intestine.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Arthritis, Experimental ; immunology ; Arthritis, Rheumatoid ; immunology ; CD4 Lymphocyte Count ; CD4-Positive T-Lymphocytes ; CD8-Positive T-Lymphocytes ; Female ; Intestinal Mucosa ; immunology ; Medicine, Chinese Traditional ; Rats ; Rats, Wistar ; Scorpions ; chemistry ; T-Lymphocyte Subsets ; drug effects ; immunology

OBJECTIVETo investigate the immunoregulatory activities of polysaccharopeptide and astragalus polysaccharides on EAC tumor-bearing mice.

METHODEhrlich's ascites carcinoma (EAC) Kunming (KM) mice were used to establish the animal model for solid tumor. Mice were randomly divided into six groups (n = 10): NS group (NS, 10 mL x kg(-1) x d(-1)), AMD group (AMD, 4 mg x kg(-1) x d(-1), 0.2 mL, only for the first 3 days), PSP group (PSP, 250 mg x kg(-1) x d(-1), 0.2 mL), APS group (APS, 250 mg x kg(-1) x d(-1), 0.2 mL), complex prescription group (PSP + APS, 250 mg x kg(-1) x d(-1), 0.1 mL) and combined treat group (AMD + PSP + APS, same dosage as above). After thirty days of treatment, immunocytochemical method was employed to detect the changes of T-lymphocyte subsets in the PBMC of tumor-bearing mice. Subsequently, the organ indexes and tumor inhibition rate were calculated and compared with those of control group.

RESULTPercentage of CD3+, CD4+ T-cell and the ratio of CD4+/CD8+ were obviously prominence in the PSP and PSP + APS groups compared with those of NS group (0.05), percentage of CD8+ T-cell was significantly decreased compared with that of AMD group; percentage of CD3+, CD4+ T-cell were obviously increased in AMD + PSP + APS group relative to that of AMD group; the thymus index of AMD group was significantly decreased compared with that of NS group, but the thymus index of AMD + PSP + APS group was obviously increased compared with that of AMD group; the weight of tumor in each administration group was significantly decreased compared with that of NS group.

CONCLUSIONPSP and PSP + APS complex prescription showed the remarkable immunoregulation on EAC mice with chemotherapy or not.

Animals ; Antineoplastic Combined Chemotherapy Protocols ; Astragalus Plant ; chemistry ; immunology ; Carcinoma, Ehrlich Tumor ; drug therapy ; immunology ; pathology ; Female ; Mice ; Polysaccharides ; administration & dosage ; immunology ; pharmacology ; therapeutic use ; Proteoglycans ; administration & dosage ; immunology ; pharmacology ; therapeutic use ; T-Lymphocyte Subsets ; drug effects ; immunology

OBJECTIVE: To explore the immunoregulatory effect of Huangqi Fuzhengtang on immunosuppressive mice. METHODS: The immunosuppressive mouse model was established by using hydrocortisone. Huangqi Fuzhengtang and Yupingfeng San apozema were given to the mice intragastrically. The contents of hemolysin, IL-2, and INF-gamma in the mouse serum and the expression of CD3(+), CD4(+), and CD8(+) in the peripheral blood lymphocytes were measured. RESULTS: Huangqi Fuzhengtang could obviously elevate the contents of hemolysin, IL-2, INF-gamma and the ratio of CD4(+) and CD8+, which was more effective than Yupingfeng San. CONCLUSION Huangqi Fuzhengtang could improve the immune function in the immunosuppressive mice.
Adjuvants, Immunologic ; pharmacology ; Animals ; Astragalus propinquus ; chemistry ; Drugs, Chinese Herbal ; pharmacology ; Hydrocortisone ; Immunocompromised Host ; drug effects ; immunology ; Male ; Mice ; Random Allocation ; T-Lymphocyte Subsets ; drug effects

OBJECTIVETo study the protective effect of Sarcandra glabra extract (SGE) on immune system in restrained mice.

METHODThe male C57BL/6 mice were randomly divided into normal control group, stress control group, 125, 500 mg x kg(-1) SGE group. The spleen lymphocyte suspensions of each group were prepared. The parameters of spleen T cells subsets, NK cell and NKT cell proportion and number was detected by Flow cytometry.

RESULTSGE regulated the balance of T cell subsets, increased the percent of NK cells and NKT cell proportion and number in restrained mice.

CONCLUSIONSGE has immunologic protective effect in restrained mice probably via the amelioration of immune cells proportion and number.

Animals ; Drugs, Chinese Herbal ; pharmacology ; Killer Cells, Natural ; drug effects ; immunology ; Magnoliopsida ; chemistry ; Male ; Mice ; Mice, Inbred C57BL ; Natural Killer T-Cells ; drug effects ; immunology ; Restraint, Physical ; Stress, Psychological ; immunology ; T-Lymphocyte Subsets ; drug effects ; immunology

OBJECTIVETo explore the influence of rosa roxburghii tratt preparation on immune function of arseniasis patients caused by burning coal.

METHODSAccording to the diagnosis standard for endemic arseniasis(WS/T 211-2001), 62 cases of arseniasis patients who resided in endemic arseniasis area in Guizhou province were selected and divided stratified randomly into two groups: rosa roxburghii tratt juice treatment group and superoxide dismutase(SOD)-enriched rosa roxburghii tratt juice treatment group, with 31 patients in each group.Each patient took 120 ml/d rosa roxburghii tratt preparation or SOD-enriched rosa roxburghii tratt orally for one month. Another 30 healthy residents from a neighbour township 12 km away where arsenic was not prevalent were selected as controls. A 2 ml blood and 50 ml urine samples were collected from individuals and the urine arsenic contents, peripheral blood T-lymphocyte subsets (CD3(+), CD4(+), CD8(+) T cell), serum immunoglobulin (IgG, IgM, IgA) and complement (C3, C4) were detected. The differences between more than two groups on above indicators were compared. The correlations between urinary arsenic and immune parameters were analyzed.

RESULTSAmong the rosa roxburghii tratt juice group, SOD-enriched rosa roxburghii tratt juice before intervention group and the control group, the levels of urine arsenic were (76.55 ± 23.02) , (72.60 ± 25.91) and (26.33 ± 11.30) µg/g Cr respectively and IgG were (11.31 ± 1.68), (11.35 ± 1.94) and (9.23 ± 1.75) g/L respectively. The differences were statistically significant(F values were 82.01, 13.82, both P values < 0.05). After intervention with rosa roxburghii tratt preparation, the levels of urine arsenic were (53.21 ± 16.51) and (51.72 ± 17.70)µg/g Cr, both decreased than before intervention (t values were 5.80 and 3.78, both P values < 0.05). The levels of CD3(+) were (44.47 ± 7.14)%, (43.44 ± 6.61)% and (70.78 ± 5.26)%, CD4(+) were (29.87 ± 5.67)%, (29.42 ± 5.87)% and (46.08 ± 5.87)%, CD4(+)/CD8(+) were(1.25 ± 0.42), (1.22 ± 0.39) and (1.79 ± 0.26) and C4 were (0.13 ± 0.08), (0.13 ± 0.09) and (0.20 ± 0.11) g/L respectively among the two treatment group before intervention and the control group. The differences were significant (F values were 178.04, 76.71, 23.13 and 5.26, all P values < 0.05). After intervention, the levels of CD3(+) were (59.73 ± 7.38)% and (66.31 ± 7.57)%, CD4(+) were (34.00 ± 7.97)% and (39.11 ± 5.81)%, CD4(+)/CD8(+) were (1.41 ± 0.37) and(1.58 ± 0.26), all increased than before intervention(t values were 12.47, 25.18, 5.41, 10.47, 3.22 and 5.05, all P values < 0.05). The levels of urine arsenic and CD3(+), CD4(+), CD4(+)/CD8(+), C4 were inversely correlated correlation, while positive correlation existed between the level of urine arsenic and IgG(r values were -0.68, -0.56, -0.51, -0.43 and 0.36, all P values < 0.01).

CONCLUSIONSThe level of urinary arsenic level is closely related to immune function suppression in arseniasis patients caused by burning coal, rosa roxburghii tratt preparation can effectively improve immune function of arseniasis patients.

Adult ; Arsenic ; urine ; Arsenic Poisoning ; etiology ; immunology ; China ; Coal ; Complement System Proteins ; immunology ; Female ; Humans ; Immunoglobulins ; immunology ; Male ; Middle Aged ; Plant Extracts ; pharmacology ; Rosa ; chemistry ; Superoxide Dismutase ; pharmacology ; T-Lymphocyte Subsets ; immunology

OBJECTIVETo observe the efficacy and safety of total glucosides of paeony capsule (TGPC) in patients with mild and moderate alopecia areata.

METHODSA total of 86 outpatients were randomly allocated into two groups of TGPC (treatment, 44 cases) and compound glycyrrhizin tablet (control, 42 cases). The treatment group was given oral TGPC, three times daily and 600 mg per time; the control group was given oral compound glycyrrhizin tablets, three times daily and 50 mg per time. In addition, both groups were given 10 mg of vitamin B(2) and tapped the bold patches with massage. The treatment course was three months for both groups. Peripheral blood T-cell subsets (CD3(+)CD4(+), CD3(+)CD8(+), Th, Ts, Th/Ts) of 10 patients randomly selected from each group respectively were tested before and after three months of treatment. The effectiveness and adverse reaction of all cases were observed each month. The safety was evaluated according to the incidence rate of adverse reaction.

RESULTSIn the treatment group, the cured and markedly effective rate was 36.36% (16/44), 50.00% (22/44) and 68.18% (30/44) at the end of first, second and third month of treatment, respectively, and the incidence rate of adverse reaction was 13.64% (6/44). In the control group, the cured and markedly effective rate was 38.10% (16/42), 57.14% (24/42) and 71.43% (30/42), respectively, and the incidence rate of adverse reaction was 16.67% (7/42). The cured and markedly effective rate and the incidence rate of adverse reaction were similar in both groups (P>0.05). TGPC and compound glycyrrhizin tablet can inhibit CD3(+)CD4(+) and CD3(+)CD8(+), and decrease the ratio of Th/Ts (P<0.05).

CONCLUSIONTGPC is effective and safe in the treatment of alopecia areata.

Adult ; Alopecia Areata ; drug therapy ; immunology ; Capsules ; Female ; Glucosides ; adverse effects ; therapeutic use ; Glycyrrhizic Acid ; adverse effects ; therapeutic use ; Humans ; Lymphocyte Subsets ; immunology ; Male ; Middle Aged ; Paeonia ; chemistry ; T-Lymphocytes ; immunology ; Tablets ; Treatment Outcome ; Young Adult

The present study was conducted in order to investigate the immunologic alterations alongside the numerical changes in peripheral blood lymphocytes(PBL) and their subsets in stomach cancer patients. Lymphocyte surface markers were determined in 85 stomach cancer patients and 49 controls by indirect immunofluorescence technique using monoclonal antibodies. Monoclonal antibodies used were Leu 2a(CD8, suppressor/cytotoxic T cells), Leu 3a(CD4, inducer/helper T cells), Leu 4(CD3, pan T reagent), Leu 11(CD16, natural killer cells) and Leu 12(CD19, B cells). The numbers of PBL, CD3+, CD4+, CD8+, CD16+ and CD19+ cells significantly decreased and the CD4: CD8 value increased in 85 patients with stomach cancer compared to those in controls(p < 0.01). In stage I(n = 17), neither PBL, their subsets nor the CD4: CD8 value were significantly different from those of the controls. In stage II(n = 17), the numbers of PBL, CD3+, CD4+ and CD8+ cells decreased(p < 0.01). In stage III(n = 24) and IV(n = 27), PBL and all subsets measured decreased(p < 0.01). The CD4: CD8 value showed significant increases in stages III and IV(p < 0.01), because the CD8+ cells decreased to a greater extent than did the CD4+ cells. The results demonstrating that the lymphocyte subsets are depressed differentially with the stage suggest that host immunity is impaired with the progression of stomach cancer.
Adult ; Age Distribution ; Aged ; Antigens, CD19/analysis ; Antigens, CD3/analysis ; CD4-Positive T-Lymphocytes/cytology ; CD8-Positive T-Lymphocytes/cytology ; Female ; Human ; Lymphocyte Count ; Lymphocyte Subsets/chemistry/*immunology ; Male ; Middle Age ; Receptors, IgG/analysis ; Sex Distribution ; Stomach Neoplasms/blood/*immunologym

OBJECTIVETo investigate the effect and mechanisms of Compound Astragalus Recipe (CAR) for regulating cellular immune in patients with myasthenia gravis (MG).

METHODSSixty MG patients were equally assigned to two groups randomly, the test group administered with CAR and the control group with prednisone for 3 months. Changes of patients' symptoms and adverse reactions were observed. The peripheral lymphocyte subsets distribution was examined by flow cytometry, and the levels of immunoglobulins and complements in the peripheral blood were measured by immuno-turbidimetry before and after treatment.

RESULTSThe total effective rate in the test group after 12-week treatment reached 80% (24/30), while that of the control group reached 83.3% (25/30), difference between them showed no statistical significance (P > 0.05). CD4+ and CD4+/CD8+ ratio were lowered significantly in both groups, but the decrement of CD4+/CD8+ ratio in the test group was more significant than that in the control group, showing significance between groups (P < 0.05). While CD8+ in the test group after treatment was significantly increased as compared with that before treatment (P < 0.05), but with no significant difference in comparing with that in the control group (P > 0.05). Serum levels of IgM and IgA in MG patients were significantly higher than normal range (P < 0.01). Levels of C3 and C4 were significantly increased in both groups after treatment (P < 0.05). Moderate high level of ALT and AST revealed transiently at the 2nd week in 5 patients of the control group, while no adverse reaction was found in the test group.

CONCLUSIONOne of the mechanisms for CAR in playing its immuno-modulate effect may be its regulation on lymphocyte subsets distribution and humoral immune function.

Adolescent ; Adult ; Aged ; Astragalus membranaceus ; chemistry ; Child ; Complement System Proteins ; metabolism ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Humans ; Immunoglobulins ; blood ; Lymphocyte Subsets ; drug effects ; Male ; Middle Aged ; Myasthenia Gravis ; drug therapy ; immunology ; Phytotherapy ; Young Adult