Role of group 2 innate lymphoid cells in mice with atopic dermatitis-like inflammatory response induced by MC903 / 中华皮肤科杂志

Objective:To explore the role of group 2 innate lymphoid cells (ILC2) in atopic dermatitis (AD) .Methods:C57BL/6J and Rag1 -/- mice served as research objects. The C57BL/6J mice were divided into 2 groups: model group topically treated with calcipotriol (MC903) on both ears every day for 14 consecutive days, control group topically treated with anhydrous ethanol alone at the same time. On day 15, peripheral blood samples were collected from the mice. After the sacrifice, the ear skin tissues were obtained for histopathological examination, and the spleens were resected. Real-time fluorescence-based quantitative PCR was performed to determine the expression of inflammatory factors in the skin and spleen tissues, and flow cytometry to determine the proportion of ILC2 in the skin tissues. The Rag1 -/- mice were divided into model group, control group and experimental group: the Rag1 -/- mice in the model group and control group received the same treatment and evaluation as the C57BL/6J mice; two days before the topical treatment with MC903, the Rag1 -/- mice in the experimental group started to be intraperitoneally injected with the monoclonal antibody CD90.2 at a dose of 300 μg/150 μl once every other 2 days for 7 sessions, with the purpose of antagonizing the function of ILC2, and other treatments were the same as those in the model group. Skin manifestations were observed, and histopathological features were evaluated. Two-independent-sample t test was used for comparisons between 2 groups, and one-way analysis of variance for comparisons among multiple groups. Results:In the model group, the ear skin of the C57BL/6J mice was apparently red, swollen and dry with crusts, and hematoxylin and eosin (HE) staining showed increased thickness of the epidermis and dermal infiltration of eosinophils; the serum level of IgE (6 751.016 ± 282.324 μg/L) was significantly higher in the model group than in the control group (6 387.038 ± 267.853 μg/L, P= 0.007) , so were the expression of interleukin (IL) -4, IL-13 and interferon (IFN) -γ in the skin tissues ( P= 0.005, 0.012, < 0.001, respectively) , but there was no significant difference in IL-5 expression ( P= 0.190) ; the expression of IL-4, IL-13 and IFN-γ in the spleen was significantly higher in the model group than in the control group (all P < 0.001) , but there was no significant difference in IFN-γ expression ( P= 0.278) ; moreover, the model group showed significantly increased proportion of ILC2 (5.604% ± 2.105%) compared with the control group (1.750% ± 1.104%, P= 0.003) . In the Rag1 -/- mice, the ear skin was obviously red, swelling and dry with crusts in the model group, and HE staining showed increased epidermal thickness and eosinophil infiltration in the dermis; the model group showed significantly increased expression of IL-4, IL-5, thymic stromal lymphopoietin (TSLP) and IL-33 in skin tissues ( P= 0.010, 0.043, 0.034, 0.007, respectively) , but no significant difference in the expression of IL-13 or IFN-γ ( P= 0.274, 0.697, respectively) compared with the control group; the proportion of ILC2 was significantly higher in the model group (5.165% ± 2.436%) than in the control group (0.835% ± 0.578%, P= 0.014) ; the experimental group showed markedly attenuated skin lesions, reduced epidermal thickness and number of eosinophils infiltrating in the dermis, but no significant difference in the expression of IL-4, IL-5, IL-13, TSLP or IL-33 compared with the model group (all P > 0.05) . Conclusion:ILC2 play a role in the mice with AD-like inflammatory response induced by MC903, which dose not depend on adaptive immunity.

Investigation of metabolism of HN-1 isolated from Millettia pachyloba in vivo and in vitro / 中国中药杂志

Ultra-fast performance liquid chromatography-mass spectrometry( UFLC-MS/MS) was used to study the anti-inflammatory active ingredient of Millettia pachyloba,6-methoxy-8,8-dimethyl-3-( 2,4,5-trimethoxyphenyl)-4 H,8 H-pyrano[2,3-f]chromen-4-one( HN-1),in liver microsomes of rats,mice,rhesus monkeys,Beagle dogs and humans metabolic stability,and compare the metabolic differences between different species. The metabolic phenotype in human liver microsomes was determined by chemical inhibitor method. Using UPLC-Q-TOF-MS/MS detection method,the in vitro metabolites of various liver microsomes were preliminarily inferred by comparing the samples incubated for 0 min and 60 min in vitro. The metabolites of HN-1 in SD rats were presumed by comparing feces,urine,plasma blanks and samples after administration. The results showed that the metabolism of HN-1 in various liver microsomes was stable,and the metabolic properties of dog and human liver microsomes were the closest. It is mainly catabolized by CYP1 A1,CYP2 D6 and CYP3 A4 isoenzymes in human liver microsomes. The metabolites of HN-1 in vitro and in vivo,including 3 in vitro metabolites and5 in vivo metabolites,were preliminarily estimated. The results laid the foundation for further pharmacological studies of HN-1.

Killing effect of Huaier combined with DC-CIK on nude mice bearing colon cancer HT29 stem cells / 中国中药杂志

To compare the therapeutic effects of different treatment methods on the nude mice bearing colon cancer HT29 cells. BalB/C nude mice colon cancer stem cell models were established and randomly divided into the following four groups, with 8 nude mice in each group: blank control group, DC-CIK group, Huaier group, and Huaier combined with DC-CIK group (combined treatment group). The mice in DC-CIK group and combined treatment group received 1×10⁶ DC-CIK cells treatment by tail vein injectionafter the tumor stem cells were inoculated for 4 days,2 times a week for three weeks. The mice in Huaier group and combined treatment group received intragastric administration at the dose of 20 g/60 kg body weight, 0.2 mL/time, once a day for a total of three weeks. The mice in control group received equal volume of normal saline. Tumor size and body weight of nude mice were measured every 2 days during treatment for three weeks in each group. After the treatment, the nude mice were sacrificed to measure the tumor weight and the tumor inhibition rate was calculated. The RT-PCR method was used to detect the expression levels of the key genes in the signal pathway. After the end of the treatment, the quality of the tumor in the Huaier group, DC-CIK group and combined treatment group was significantly lower than that in the control group; the quality in combined treatment group was significantly lower than that in Huaier group and DC-CIK group.Among them, the tumor inhibition rate reached 46.77% in the combined treatment group. In respect of changes in expression levels of key genes in the signaling pathway, the mRNA expression levels of key genes PI3KR1 and Akt in PI3K/Akt pathway, key genes Wnt1 and CTTNB1 in Wnt/-catenin pathway, and key genes Notch1, Notch2, Notch3 in Notch pathway in the combined treatment group were lower than those in DC-CIK group and Huaier group. The Huaier combined with DC-CIK group showed best therapeutic effect among different treatment methods for HT29 stemcell colon tumors in nude mice, providing a new idea for clinical treatment of colon cancer.

Optimization of Water Extraction Technology of Yangxue Jiegu Capsule by Orthogonal Test / 中国药房

OBJECTIVE:To optimize the water extraction technology of Yangxue Jiegu capsule. METHODS:Using adding amount of water,decoction time and decoction times as investigation factors,ferulic acid content,icariin content and yield rate as investigation indexes,orthogonal test was used to optimize the optimal water extraction technology of Yangxue Jiegu capsule;and verification test was conducted. RESULTS:The optimized extraction technology was 10-fold water,extracting 3 times,1 h once. Contents of ferulic acid and icariin were 0.38,1.23 mg/g(n=3),yield rate was 28.2%(n=3),average comprehensive score was 97.11 (RSD=2.77%,n=3). CONCLUSIONS:Optimized extraction technology is feasible,accurate,reliable,and can be used for the extraction and preparation of Yangxue Jiegu capsule.

Expression of CXCR3 and CCR5 chemokine receptor in spleens of patients with primary immune thrombocytopenia / 中华血液学杂志

<p><b>OBJECTIVE</b>To study CXCR3 and CCR5 chemokine receptor expression in spleens of patients with primary immune thrombocytopenia (ITP) and its clinical significance.</p><p><b>METHODS</b>The splenectomy specimens from 10 ITP patients (ITP group) and 8 patients with traumatic splenic rupture (normal control group) were studied. Immunohistochemistry (IHC) was used to study the positive rate of CXCR3 and CCR5. Western blot was performed to detect CXCR3 and CCR5 protein expression, while real-time polymerase chain reaction (RT-PCR) was conducted to analyze their mRNA expression.</p><p><b>RESULTS</b>The positive rate of CXCR3 and CCR5 were both higher in ITP group (90% and 100%, respectively) than those in control group (75% and 87.5%, respectively)(P < 0.05). The differences were statistically significant (P < 0.05). Protein and mRNA level of CXCR3 in ITP group were 3.0 and 3.5 times as high as those in control group, respectively. Those of CCR5 in ITP group were 1.2 and 1.7 times as high as those in control group, respectively.</p><p><b>CONCLUSION</b>High expression of CXCR3 and CCR5 may play a part in the splenic immune disorders in patients with ITP.</p>

In vitro activation of bone marrow natural killer T cells of aplastic anemia patients / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the quantitative and qualitative changes of TCRVα24(+)Vβ11(+) natural killer T (NKT) cells from bone marrow (BM) of aplastic anemia (AA) after in vitro stimulation of α-galactosylceramide (α-Galcer).</p><p><b>METHODS</b>NKT cells in the bone marrow mononuclear cells (BMMNCs) from either AA patients or healthy controls were enumerated with flow cytometry. BMMNCs were cultured in RPMI1640 medium supplemented with either α-Galcer and rhIL-2 or α-Galcer, rhIL-2 and rhG-CSF. The proliferative capacity of NKT cells was determined by NKT cell numbers before and after in vitro culture. Expression of intracellular IFNγ and IL-4 in activated NKT cells was analyzed with flow cytometry.</p><p><b>RESULTS</b>In AA group, the percentage of NKT cells in BMMNCs was (0.19 ± 0.09)%. Addition of rhG-CSF into the α-Galcer/rhIL-2 culture medium resulted in significantly reduced expansion of NKT cells (67.45 ± 29.42-fold vs 79.91 ± 40.56 fold, P < 0.05). Meanwhile, addition of rhG-CSF reduced IFNγ positive NKT cells \[(37.45 ± 7.89)% vs (62.31 ± 14.67)%, P < 0.01\] and increased IL-4 positive NKT cells \[(55.11 ± 12.13)% vs (27.03 ± 9.88)%, P < 0.01\]. In healthy control group, the percentage of NKT cells in BMMNCs was (0.25 ± 0.12)%. Addition of rhG-CSF into the α-Galcer/rhIL-2 culture medium also significantly reduced expansion of NKT cells (97.91 ± 53.22-fold vs 119.58 ± 60.49-fold, P < 0.05), reduced IFNγ positive NKT cells \[(28.65 ± 10.63)% vs (50.87 ± 12.66)%, P < 0.01\], and increased IL-4 positive NKT cells \[(66.53 ± 14.96)% vs (31.11 ± 10.07)%, P < 0.01\].</p><p><b>CONCLUSION</b>Compared to those from healthy controls, BMMNCs from AA patiants have a reduced fraction of NKT cells, which possesses a decreased potential to expand in vitro in response to α-Galcer stimulation, and produce more IFNγ(+) NKT1 cells. rhG-CSF, in combination with α-Galcer, confers polarization of NKT cells towards IL-4(+) NKT2 subpopulation.</p>

Cyclic adenosine monophosphate signal pathway in targeted therapy of lymphoma / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To review the role of cyclic adenosine monophosphate (cAMP) signal pathway in the pathogenesis of lymphoma and explore a potential lymphoma therapy targeted on this signaling pathway.</p><p><b>DATA SOURCES</b>The data cited in this review were mainly obtained from the articles listed in Medline and PubMed, published from January 1995 to June 2009. The search terms were "cAMP" and "lymphoma".</p><p><b>STUDY SELECTION</b>Articles regarding the role of the cAMP pathway in apoptosis of lymphoma and associated cells and its potential role in targeted therapy of lymphoma.</p><p><b>RESULTS</b>In the transformation of lymphocytic malignancies, several signal pathways are involved. Among of them, the cAMP pathway has attracted increasing attention because of its apoptosis-inducing role in several lymphoma cells. cAMP pathway impairment is found to influence the prognosis of lymphoma. Targeted therapy to the cAMP pathway seems to be a new direction for lymphoma treatment, aiming at restoring the cAMP function.</p><p><b>CONCLUSIONS</b>cAMP signal pathway has different effects on various lymphoma cells. cAMP analogues and phosphodiesterase 4B (PDE4B) inhibitors have potential clinical significance. However, many challenges remain in understanding the various roles of such agents.</p>

Cutaneous nerve morphology and protease activated receptor 2 expression in pruritic skin lesions of atopic dermatitis / 中华皮肤科杂志

Objective To study the role of cutaneous nerve and protease activated receptor 2 (PAR2)in the development of pruritus in atopic dermatitis(AD).Methods Dermal sheets were prepared from chronically pruritic skin lesions of 7 patients with AD,as well as from the normal skin of 7 healthy human controls.Double labeled immunofluorescence was performed using mouse anti-protein gene product 9.5(PGP9.5)monoclonal antibody and rabbit anti-substance P(SP)polyclonal antibody to observe the morphological changes in cutaneous nerve fibers,and Image-Pro Plus 6 software was used to semiquantitively assess the length,diameter of nerve fibers,integral optimal density of PAR2 and SP in dermal sheets.Results Immunofluoresence double staining showed that PAR2 co-expressed with PGP9.5 or SP in cutaneous nerve fibers.Compared with the normal control skin,both the total length and average diameter of PGP 9.5-expressing nerve fibers were increased(11051.8±1900.9 μm vs 7264.0±2659.9 μm,4.23±0.15 μm vs 3.95±0.15 μm,both P＜0.01)in pruritic lesions,while only the average diameter of SP-expressing nerve fibers was up-regulated(3.99±0.20 μm vs 3.80±0.07 μm,P＜0.05),and the total length of them remained unchanged(4304.7±1455.0 μm vs 3380.0±1735.4 μm,P＞0.05).Also,increased integral optimal density was observed for SP and PAR in pruritic lesions in comparison with the normal control skin (27.71±16.52 vs 12.63±4.31.35.99±8.63 vs 22.69±9.56.both P＜0.05).Conclusion Our results indicate a hyper-plasia of cutaneous nerve fibers in chronic itchv skin lesions of AD and an increase in the expression of PAR2 and SP in the cutaneous nerve fibers,suggesting that the signal enhancement in PAR2 pathway may be related to the mechanism of pruritus in patients with AD.

Effects of PML-RARalpha on cAMP-induced AML cell differentiation / 中国实验血液学杂志

To explore the molecular mechanisms of acute promyelocytic leukemia cell differentiation induced by cAMP combined with low-dose As2O3, the PR9 cell line, which was stably transfected by PML-RARa fusion gene, was used as in vitro model. The effects of PML-RARa on cAMP-induced AML cell differentiation were evaluated according to cell growth, cell morphology, cell surface antigen as well as luciferase reporter gene assay, in the cells before and after the treatment with cAMP and/or As2O3. The results showed that cAMP alone could slightly increase the expression of CD11b in the PR9 cells expressing the PML-RARa fusion protein, but could not induce these cells to differentiate. The cells presented the terminal differentiation morphology and significantly increased CD11b expression only under the treatment of cAMP combined with As2O3. In addition, PML-RARa had strong inhibitory activity on the transcription of the reporter gene containing cAMP response elements. In conclusions, the PML-RARa fusion protein could dramatically block the signaling pathway of cAMP during the AML cell differentiation.

Expression of specific antibodies against platelet glycoproteins in patients with mds and its significance / 中国实验血液学杂志

The aim of this study was to find platelet specific autoantibodies against glycoproteins in myelodysplastic syndrome (MDS) and to explore its role in pathogenesis of MDS. The plasma autoantibodies against GP IIb/IIIa and GP Ib/IX were measured by using a modified monoclonal antibody specific immobolization platelet antigens assay (MAIPA). Absorbance greater than mean value plus tripled standard deviation recorded from the normal controls were regarded as positive. The results indicated that the total positive rate in patients with MDS was 16.67% (5/30), the total positive rate in patients with ITP was 46.67% (14/30), the difference between MDS group and ITP group was significant (P < 0.05). It is concluded that partial patients with MDS have plasma specific autoantibodies against platelet GP II b/III a and GP Ib/IX, indicating correlation of thrombocytopenia of patients with immune factors and the autoantibody-mediated platelet destruction may be involved in the pathogenesis of MDS. It provides a new basis for immunosuppression therapy for MDS.

Activation of adenylate cyclase influences the sensitivity of acute promyelocytic leukemia cell lines to ATRA / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the molecular mechanism of APL cell resistance to ATRA.</p><p><b>METHODS</b>The ATRA sensitive and resistant APL cell lines, NB4 and NB4-R1, were used as in vitro models. The effects of specific inhibitors and activators of adenylate cyclase (AC) and phosphodiesterase (PDE) on ATRA-induced differentiation was evaluated by cell morphology, cell surface antigen expression and nitroblue-tetrazolium (NBT) reduction assays.</p><p><b>RESULTS</b>SQ22536, a specific antagonist of AC, could dramatically block ATRA-induced NB4 cell differentiation. When ATRA + SQ22536 group compared with ATRA group, the positivity of CD11b decreased from (95.9 +/- 2.5)% to (60.3 +/- 7.1)%, while the A(540) in NBT reduction assay decreased from 0.585 +/- 0.092 to 0.170 +/- 0.028 (P < 0.05). Forskolin, an agonist of AC, could overcome the resistance of NB4-R1 cells to ATRA. When ATRA + forskolin group compared with ATRA group, the positivity of CD11b increased from (34.3 +/- 5.3)% to (94.6 +/- 2.4)%, while the A(540) in NBT reduction assay increased from 0.110 +/- 0.028 to 0.395 +/- 0.049 (P < 0.05). In contrast, the specific antagonist and agonist of PDE, 3-isobutyl-1-methylxanthine (IBMX) and calmodulin, exerted little impact on ATRA treatment.</p><p><b>CONCLUSIONS</b>The defaults in the initiation of AC activation may contribute to the resistance to ATRA in some APL cells.</p>

Expression of nuclear factor ?B and the effect of topical tacrolimus ointment on lesional atopic dermatitis skin / 北京大学学报(医学版)

Objective: To investigate the role of nuclear factor ?B (Rel/NF-?B) in pathogenesis of atopic dermatitis(AD) and the effect of topical 0.1%(mass fraction) or 0.03%(mass fraction) tacrolimus ointment on expression of NF-?B in lesional AD skin. Methods: Immunohistochemistry has been employed to study the expression of NF-?B in normal skin and lesional AD skin before and after using topical tacrolimus ointment. Results: The expressions of NF-?Bp50 and NF-?Bp65 were scattering or negative in normal keratinocytes. NF-?Bp50 was overexpressed on nuclear of basal and suprabasal keratinocytes in 9 cases of AD, NF-?Bp65 was overexpressed in cytoplasm and perinuclear of basal and suprabasal keratinocytes. After using topical tacrolimus ointment for three weeks , nuclear NF-?Bp50 expressed on basal and suprabasal keratinocytes were lost and NF-?Bp50 was expressed sparsely on basal keratinocytes cytoplasm or nuclear. NF-?Bp65 was expressed sparsely on basal and suprabasal keratinocytes cytoplasm. Conclusion: These data suggest that increased NF-?B activity may represent the basis of initiation or maintenance of the skin inflammatory response in atopic dermatitis. Topical tacrolimus may directly or indirectly inhibit NF-?B nuclear expression in keratinocytes and inhibit skin innate immuno-inflammatory response in atopic dermatitis that related to NF-?B.

Efficacy and Safety of TacroUmus Ointment for the Treatment of Atopic Dermatitis in Chinese Children / 中华皮肤科杂志

Objective To evaluate the efficacy and safety of 0.03% tacrolimus ointment for the treatment of atopic dermatitis (AD) in Chinese children. Methods A total of 139 children, 2 to17 years of age, with moderate to severe AD from 5 study centres were enrolled in this multicentre, randomized, double-blind, vehicle-controlled, parallel group study. Treatment with 0.03% tacrolimus ointment or vehicle was applied twice daily to the affected areas for 3 weeks. Visits were scheduled on day 1 (base line, before treatment) and 1, 2, 3 weeks after the treatment. The main therapeutic parameter was the efficacy rate at the end of the treatment. Results The efficacy rates were 84.6% and 29.0% for tacrolimus group and vehicle group, respectively (P

Efficacy and Safety of Tacrolimus Ointment for the Treatment of Atopic Dermatitis in Adults / 中华皮肤科杂志

Objective To observe the clinical efficacy and safety of 0.1% and 0.03% tacrolimus ointment in patients aged 18 to 65 years with atopic dermatitis (AD). Methods Treatment was given twice daily to all affected areas for 3 weeks in a multicentre, randomized, double blind, parallel, and vehicle-controlled study. Follow-up visits were scheduled on day 1(baseline), and 1, 2 and 3 weeks post-treatment. The therapeutic effect and safety were evaluated. Results A total of 211 adults with moderate to severe AD in 6 study centres were enrolled in the efficacy evaluation. The efficacy rates were 88.4%, 77.8% and 30.0% in patients treated with 0.1%, 0.03% tacrolimus ointment, and the vehicle, respectively (P

Effect of reduced glutathione on the proliferation,oxidative stress and transforming growth factor?1 expression of human hepatic stellate cells / 中华消化杂志

Objective To investigate the impact of reduced glutathione(GSH) on the prolifera- tion,oxidative stress and transforming growth factor?1(TGF-?1) expression of human hepatocytes and hepatic stellate cells(HSCs)(LX-2 cell line).Methods Human hepatocytes and HSCs were incubated with various concentrations of GSH(0.5—50 mmol/L or 0.5—10 mmol/L).The effects of GSH on the proliferation of hepatocytes and HSCs were studied by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphennyhera- zolium bromide colorimetric assay.Human hepatocytes and HSCs were co-cultured with GSH and ferric nitrilotriacetic acid,superoxide dismutase (SOD) activity and malondialdehyde (MDA) contents were detected.HSCs were incubated with high(5.0 mmol/L),media(2.5 mmol/L) and low (0.5 mmol/L) concentrations of GSH,the expressions of TGF-?1 mRNA and protein were detected by ELISA and real- time PCR.Results In concentration ranged from 2.5 to 10 mmol/L,the GSH could promote the pro- liferation of hepatocytes but no HSCs,significantly increased the activity of SOD and decrease the con- tents of MDA in hepatocytes and HSCs,and inhibited the expression of TGF-?1 in HSCs.Conclusions GSH can not only promote the proliferation of hepatocytes,but also protect hepatocytes and HSCs from oxidative stress,and inhibit the secretion of TGF-?1 in HSCs.GSH may play a role in hepatocellular protection,antioxidation and anti-fibrosis.

Effect of bilateral subotal thyroidectomy for juvenile hyperthyroidism / 实用儿科临床杂志

Objective To evaluate the effect of medicial and surgical treatment for juvenile hyperthyroidism.Methods Ortapazole was administ rated separately in drug therapy group for 1.5-2 years.Bilateral subtotal thyroidectomy was done in surgical therapy group.Results In drug therapy group,effective rate was 60 percent in 6 months and 70 percent in one year.Recurrence rate was 40 percent after drug withdrawal in 2 years curative rate was 60 percent.In surgical therapy group,the average stay in hospital was 16 days.There was no nerve injury,parathyroidal hypofunction,thyroid crisis or hypothyroidism complications,with 100 percent curative rate after 2 years′ followup.Postoperative growth and development were normal.Conclusions Surgical treatment may be suitable for those who have no response to drug therapy,with recurrence after drug withdrawal,whose compression symptom was obvious,with moderate and severe hyperthyroidism or those who could not take medicine persistently.Bilateral subtotal thyroidectomy applied in juvenile hyperthyroidism could achieve quick and better recovery,and has no influence on the juvenile growth and development.