Objective: The effects of estrogen on cellular immunity were investigated to explore the immunomodulatory properties of estrogen. Methods: In the present study, the in vitro influences of estrogen at 10~(-8),10~(-7 )and 10~(-6) mol/L on the ConA-induced proliferation of thymocytes were determined by MTT assay. Results: The results showed that estrogen at all concentrations we used suppressed the ConA-induced proliferation of thymocytes. The inhibitive effects had dose- and time- dependent pattern. Conclusion: These results suggested that estrogen had inhibiting effects on the cellular immunity.

ve To understand the effects of fenvalerate on the secretion of sex hormone and humoral im-mune function and to explore their mechanism. Methods 20 female SD rats aged 4 weeks were randomly divided into 2 groups: fenvalerate group treated by peritoneal injection with 1/10 LD50 of fenvalerate and control group treated with disinfected bean oil. The serum specimens were obtained from rats of 2 groups at the 4th week after the peri-toneal injection. The levels of interleukin-6 (IL-6). tumor necrosis factor-? (TNF-?) and immunoglobulin G (IgG) in serum were measured with ELISA. The levels of 17?-estradiol (E2) and testosterone (T) were measured with ra-dioimmune assay. Results The levels of IL-6, TNF-? and IgG in serum specimens of rats were 2.244 ng/ ml, 0.360 ng/ml and 4.928?g/ml for fenvalerate group, 2.805 ng/ml, 0.439 ng/ml and 3.825?g/ml for control group respec-tively. The levels of IL-6 and TNF-? in serum specimens of rats were significantly lower than those of control group (P 0.05. Conclusion Fenvalerate could effect the hu-moral immune system and the levels of sex hormones. Its passible mechanism was that fenvalerate could affect the levels of sex hormones first, and then the whole immune system further.

Objective To test the performance of field blood transportation vehicle from the aspects of heat preservation,refrigeration and oil consumption.Methods Two vehicles without power supply were stowed with maximum load,one with red cell suspension,and the other with frozen plasma.With the environmental temperature higher than 35 ℃ or lower than 25 ℃,related time lengths were detected respectively.Results The holding time of the vehicles was from 1.5 h to 7 h,refrigerating time was from 15 min to 138 min,and a full tank of gasoline was exhausted around 5～7 hours.Conclusion During line-haul,field blood transportation vehicle has to be equipped with additional cooling agents,in case the refrigeration equipment out of work for the reason of gasoline exhaustion.

Objective To investigate the abnormality of intrahepatic biliary epithelial cells autophagy in primary biliary cirrhosis (PBC) mice,and explore the mechanism of UC-Mesenchymal stem cell (MSCs) in treating PBC.Methods After establishing the PBC model,we divided them into the PBC model group;the UC-MSCs treatment group and the Stattic group [Signal transducer and activator of transcription 3 (STAT3) inhibitor group].Six mice were used as the control group.Liver pathology and the serum pyruvate dehydrogenase complex E2 subunit (PDC-E2) antibody titers were detected.Autophagosome of the intrahepatic biliary epithelial cell was observed by electronic microscope.Protein levels of STAT3/pSTAT3,Beclin-1 were detected by western blot.We cultured human intrahepatic biliary epithelial cells in vitro,and down regulated STAT3.After stimulated by GCDC,we co-cultured them with UC-MSCs,and collected the cells in order to detect LC3 Ⅱ.The measurement data were compared with t test or single factor analysis of variance.Results Compared with the control group,periportal inflammatory cell infiltration and granuloma formation were observed in the PBC group.MSCs treatment decreased the infiltration of inflammatory cells.The level of antiPDC-E2 of the PBC group (107±18) ng/ml was higher than that of the control group (42±6) ng/ml (q=6.326,P＜0.01),MSCs treatment down regulated anti-PDC-E2 level (43±4) ng/ml (q=5.801,P＜0.01).More autophagosomes in the PBC group (5.00±1.29) than the control group (1.75±0.25) were observed (q=4.061,P＞0.05).Western blot showed that the level of Beclin-1 was higher in PBC group (1.80±0.36) than the control group (0.40±0.20) (q =6.757,P＜0.01),MSCs reduced the expression of Beclin-1 (0.86±0.06)(q=4.536,P＜0.05) as well as Stattic (0.72±0.03) (q=5.226,P＜0.05).PBC group had a higher expression level of STAT3 (1.80±0.42) (q=5.730,P＜0.05) and pSTAT3 (2.04±0.29)(q=6.492,P＜0.01) than the control group (0.50±0.05)(0.91±0.14).MSCs treatment decreased the expression of STAT3 (0.51±0.13)(q =5.703,P＜0.01) and pSTAT3 (0.76±0.07) (q =7.388,P＜0.01) in intrahepatic biliary epithelial cells.After down regulated STAT3 of HiBECs,MSCs reduced the expression of LC3 Ⅱ of HiBECs.Conclusion The intrahepatic biliary epithelial cells autophage of PBC mice is abnormal,MSCs can alleviate PBC by down regulating the autophage of intrahepatic biliary epithelial cells via STAT3.

Objective:To explore whether sera from patients with systemic lupus erythematosus affect the senescence of MSCs, and which signaling pathway might be involved in.Methods: Human sera were collected from sex-matched,age-matched active SLE patients (n=6) and healthy volunteers (n=6).Then the complement was inactivated at 56℃ for 30 min.MSCs were cultured in DMEM/F12 containing 10% SLE or normal serum.Three days later MSCs were stained with SA-β-gal.The percentages of SA-β-gal positive cells were evaluated.The mRNA levels of p53 and p21 were detected by Real time PCR.The protein levels of p53/p21 and NF-κB ( p65) were examined by Western blot.Cell proliferation was evaluated by CCK8 assay.Results: Compared with normal serum, MSCs from either healthy controls or SLE patients showed more SA-β-gal positive cells in SLE serum,as well as higher levels of p53/p21.In addition,SLE serum also inhibited the capacity of MSC proliferation.In MSCs treated with SLE serum, the level of NF-κB (p65) protein was significantly upregulated.Conclusion:SLE serum can promote the senescence of bone marrow MSCs,which might be mediated by NF-κB signaling pathway.

Objective To study the blood concentration of hydroxychloroquine in patients with systemic lupus erythematosus (SLE) treated with different doses, and analyze the relationship between blood concentration of hydroxychloroquine and disease activity, and evaluate its safety.Methods Forty SLE patients were randomly divided into 2 groups, each group contained 20 cases.The patients in group A were treated with hydroxychloroquine (0.4 g, qd), while patients in group B were treated with hydroxychloroquine (0.2 g, qd).The treatment lasted more than six months in every patient.The blood concentrations of hydroxychloro-quine were detected by high performance liquid chromatography.The clinical and laboratory indices were collected.The systemic lupus erythematosus disease activity index (SLEDAI) score was recorded.The doses and varieties of combined hormone, immunosuppressant were recorded.The correlation of blood concentration of hydroxychloroquine and disease activity was analyzed.The significance was determined by Student's t test and Pearson correlation analysis.Results In SLE patients, the average blood concentration of hydro-xychloroquine was (402±190) ng/ml in group A and (150±60) ng/ml in group B (t=7.471, P＜0.01).The disease activities of patients in the two groups showed no significant difference (t=-0.172, P＞0.05).The platelet counts of patients in group A were significantly higher than those in group B[(188±88)×109/L vs (158 ±87) ×109/L] (t=4.375, P＜0.05).However, the other laboratory parameters showed no significant difference between the two groups (P＞0.05).Conclusion The results of this study indicate that the blood concentrations of hydroxy-chloroquine are significantly different in different dosages.The high dose of hydroxy-chloroquine is related to high platelet number in lupus patients.These findings suggest that hydroxychloroquine is safe and effective for SLE patients.

Objective To explore the preventive effect of early umbilical cord mesenchymal stem cells (UC-MSCs) transplantation on MRL/lpr mice and the underly mechanisms.Methods Fourteen 10-week-old MRL/lpr mice were labeled and numbered.They were randomly divided into 2 groups by using random number table and injected with 1 ×106 UC-MSCs or PBS via tail vein respectively.Proteinuria was measured with Bradford method every 4 weeks.All mice were sacrificed at the age of 28 weeks, with the level of serum antidsDNA antibody and IL-17 detected by enzyme linked immunosorbent assay (ELISA).Splenic Th17 cells, as well as regulatory T cells (Treg) were examined by flow cytometry.Data were analyzed with t test and Pearson's correlation test.Results The onset of proteinuria was delayed for 4 weeks in UC-MSC-treated group compared with that in the control group.At the age of 28 weeks, the 24 hour proteinuria [(1.78±0.17) mg vs (4.77±0.98)mg, t=2.99, P＜0.05] and the spleen weight [(0.149±0.009) g vs (0.273±0.052) g, t=2.33, P＜0.05] in UC-MSCtreated group were significantly lower than those in the control group.There was also a trend of the decline of serum anti-dsDNA antibody and IL-17 level after UC-MSCs transplantation.Compared with those in the control group, both the percentage and the absolute number of Th17 cells were significantly decreased in UC-MSC-treated group [(0.90±0.19)％ vs (2.81±0.50)％, t=3.54, P＜0.01 and (3.7±0.8)×105 vs (19.3±3.7)×105, t=4.12,P＜0.01].Meanwhile, the percentage of Treg elevated after UC-MSCs treatment.The ratio of Th17/Treg was significantly lower in UC-MSC-treated group than that in the control group (0.11±0.03 vs 0.50±0.09, t=4.23,P＜0.01).Both the ratio of Th17/Treg (r=0.73, P＜0.01;r=0.59, P＜0.05) and serum IL-17 level (r=0.78, P＜0.01;r=0.56, P＜0.05) was positively correlated with the level of 24 hour proteinuria and anti-dsDNA antibody respectively in MRL/lpr mice.Conclusion Early UC-MSCs transplantation helps to delay disease onset and ameliorate disease progression in MRL/lpr mice, which may act through the modulation of Th17/Treg balance.

Objective Whether the bone marrow cells (BMC) derived from systemic lupus erythematosus (SLE) could transmit autoimmune disease was studied for the purpose of clarifying the role of BMC in SLE pathogenesis.The effects of bone marrow mesenchymal stem cells (MSC) from SLE and control mice on the SLE BMC-induced symptoms were compared to elucidate the role of MSC in SLE.Methods Six-week-old B6.MRL-Fas mice were randomly divided into 3 groups.One group was transplanted with BMC from the 30-week-old B6.MRL-Faslg mice.One group was co-transplanted with BMC from the 30-week-old B6.MRL-Fasr mice and bone marrow MSC from the age-matched B6.MRL-Faslpr mice.One group was co-transplanted with BMC from the 30-week-old B6.MRL-Faslg mice and bone marrow MSC from the age-matched C57BL/6 mice.Before transplantation,the recipient mice received irradiation by an X-ray source.The levels of serum antinuclear antibody (ANA) and proteinuria were measured with enzyme linked immunosorbent assay (ELISA) and Bradford method every 4 weeks,respectively.The survival rate was recorded.All mice were sacrificed 18 weeks later.Splenic plasma cells,Th1,Th2 and Th17 cells were measured by flow cytometry.Statistical analyses were performed using the independent t test and ANOVA.Results Eight weeks after transplantation,ANA was positive in all the recipient mice.However,there was no significant difference between the three groups (P＞0.05).No proteinuria was observed in all the recipient mice.The mice received BMC from the 30-week-old B6.MRL-Fasr mice and bone marrow MSC from the age-matched B6.MRL-Fasr mice showed an elevated trend of the percentages of splenic plasma cells,Th1,Th2 and Th17 cells compared with the other two groups,plasma cells [(1.05±0.16)％,(0.58±0.11)％,t=2.53,P＞0.05;(1.05±0.16)％,(0.71±0.18)％,t=1.45,P＞0.05],Th1 cells [(6.6±2.2)％,(5.7±1.0)％,t=0.38,P＞0.05;(6.6±2.2)％,(4.0±1.7)％,t=0.96,P＞0.05],Th2 cells [(3.3±0.4)％,(2.1±0.6)％,t=1.76,P＞0.05;(3.3±0.4)％,(2.2±0.6)％,t=1.51,P＞0.05],Th17 cells [(2.30±0.71)％,(1.31±0.31)％,t=1.27,P＞0.05;(2.30±0.71)％,(1.12±0.27)％,t=1.67,P＞0.05].However,there was no significant difference between the groups.The survival rate of the three groups was 43％,43％ and 80％ respectively.And the survival rate of the mice received BMC from the 30-week-old B6.MRL-Fasr mice and bone marrow MSC from the age-matched C57BL/6 mice was significantly higher than those of the other groups.Conclusion Our results indicate that BMC from SLE can transmit autoimmune disease.The bone marrow MSC can not prevent lupus-like presentations induced by BMC from SLE.Transplantation of bone marrow MSC from C57BL/6 mice can significantly elevate the survival rate.

Objective The purpose of this study is to observe the changes of immune cell subsets in lupus mice after umbilical cord mesenchymal stem cells (UC-MSCs) transplantation. Methods B6.MRL-Faslpr lupus mice were randomly divided into the following three groups: the UC-MSCs treated group, the fibroblast like synoviocytes (FLS) treated group and the untreated group. MSC (1×106) or FLS (1×106) were injected into the tail vein of lupus mice respectively. Four weeks after treatment, the spleen index was calculated. The pathological changes of kidney were assessed by HE staining. The frequencies of immune cell subsets in spleen and macrophage in kidney as well as abdominal cavity were analyzed by flow cytometry. Data were analyzed with t test. Results The spleen index of UC-MSCs treated lupus mice [(79 ±9) mg/10 g] and IgG level [(7.5±1.5) mg/ml] were significantly decreased when compared with FLS treated group [(147±23) mg/10 g, t=2.78, P<0.01] [(17.0 ±2.8) mg/ml, t=3.00, P<0.01] and the untreated group [(156 ±16) mg/10 g, t=4.29, P<0.01] [(16.7 ±1.6) mg/ml,t=4.01, P<0.01]. HE staining also showed that the pathological changes of kidney were alleviated after MSCs transplantation. In addition, the frequency of plasma cells in the untreated group [(2.61 2± 0.318)% vs (0.306±0.017)%, t=7.22, P<0.01] and the FLS treated group [(2.412±0.297)% vs (0.306±0.017)%, t=7.07, P<0.01] were markedly higher than MSCs treatment [(0.306 ±0.017)%]. Moreover, the frequency of CD25+Foxp3+/CD4+Treg in the MSCs treated group [(15.08±0.81)%] was significantly increased compared with the untreated group [(8.02 ±0.47)%, t=7.45, P<0.01] and FLS treated group [(8.80 ±0.23)%, t=7.39, P<0.01]. MSCs treatment resulted in a decrease in CXCR5+PD1+/CD4+Tfh and IFNγ+/CD4+Th1 subset, compared with the untreated group [(14.3±1.5)%vs (31.5±3.3)%, t=5.25, P<0.01] [(1.78±0.27)% vs (5.93±1.56)%, t=2.60, P<0.05] and the FLS treated group [(14.3±1.5)%vs (28.8±2.2)%, t=5.49, P<0.01] [(1.78±0.27)%vs (4.88±0.81)%, t=3.61, P<0.01]. The frequency of macrophage in kidney of the MSCs treated group [(3.52 ±0.37)%] was markedly increased compared with the untreated group[(1.58±0.29)%, t=3.25, P<0.01], while neither the IL4+/CD4+Th2 subset nor the IL17+/CD4+Th17subset and the frequency of macrophage in abdominal cavity showed significant changes in the three groups. Conclusion These findings suggest that the therapeutic effects of MSCs on lupus mice may mediate through increasing the frequency of spleen Treg and renal macrophage and decreasing the frequency of Tfh, Th1 and plasma cells.

OBJECTIVETo explore the molecular mechanism underlying the DEL phenotype among RhD negative ethnic Han individuals from Jiangsu, China.

METHODSThe DEL phenotype was determined by an adsorption elution test among 57 RhD negative blood donors. The Rh C, c, E, and e phenotypes were detected by a tube method. PCR with sequence-specific primering (PCR-SSP) assay was used to determine the RHCE genotypes. The RHD gene of the DEL individuals were amplified with polymerase chain reaction and subjected to Sanger sequencing analysis.

RESULTSAmong the 57 RhD negative donors, 10 (17.54%) were determined as having the DEL phenotype. The major RhCE phenotypes for DEL and RhD negative cases were RhCcee (80.0%) and Rhccee (61.7%), respectively. All RHD gene sequences of the 10 individuals have harbored a G>A mutation at position 1227 of exon 9.

CONCLUSIONA proportion of RhD negative individuals determined by routine serological method are actually DEL with RHD gene mutations. RHD *1227A is the most prevalent DEL genotype among ethnic Han Chinese from Jiangsu. Further research on the phenotype and underlying molecular mechanism of DEL is important for blood transfusion.

Alleles ; Asian Continental Ancestry Group ; ethnology ; genetics ; Base Sequence ; Blood Donors ; China ; ethnology ; Exons ; Genotype ; Humans ; Male ; Molecular Sequence Data ; Phenotype ; Polymorphism, Genetic ; Rh-Hr Blood-Group System ; genetics