Objective To investigate the regulatory effect of glutamate signaling pathway on filopodia formation in epidermal cells and on melanosome transfer.Methods Epidermal melanocytes and keratinocytes were isolated from human foreskin and subjected to subculture.After two to three passages of subculture,the melanocytes and keratinocytes were cultured alone or in combination with or without the presence of MK801 (an antagonist of N-methyl-D-aspartic acid (NMDA) receptor) of 100 μmol/L,or NMDA (the activator of NMDA receptor) of 100 μmol/L,for 24 hours.The melanocytes irradiated with UVB at 311 nm served as the control.Scanning electron microscopy was used to observe the appearance of filopodia and dendrites of melanocytes and keratinocytes.Melanosome transfer was visualized under confocal laser scanning microscopy after double immunofluorescent staining.Results Although no obvious changes were observed in the number of dendrites in monocultured melanocytes after treatment with MK801 or NMDA for 24 hours,dendrites became thinner at the terminus and longer with a decrease in the number and length of filopodia after MK801 treatment,but thicker and shorter with an increase in the number and length of filopodia after NMDA treatment compared with untreated monocultured melanocytes.In the coculture system,filopodia were observed between the untreated melanocytes and keratinocytes,and the number of filopodia in melanocytes was larger in the side adjacent to keratinocytes than in the opposite side.Compared with the untreated coculture system,the number of both filopodia connecting melanocytes and keratinocytes and filopodia extending from melanocytes to keratinocytes decreased in the coculture system after treatment with MK801 of 100 μmol/L,but increased after treatment with NMDA of 100 μmol/L,for 24 hours.Melanosomes were found in keratinocytes cocultured with melanocytes without treatment,which were decreased in number after 24-hour treatment with MK801 of 100 μmol/L,but increased in number and even present in keratinocytes nonadiacent to melanocytes after 24-hour treatment with NMDA of 100 μmol/L.Conclusion Glutamate signaling pathway may modulate the transfer of melanosomes from melanocytes to keratinocytes via modulating the appearance of melanocyte dendrites and formation of filopodia.

Objective T o observe the therapeutic effect of Radix Astragali Injectio on psoriasis and it s possible pharmaceutical mechanism. Methods Two groups of patients with psoriasis vulgaris were treated with either comprehensive regi men or comprehensive regimen plus Radix Astragali Injectio. The clinical respons es in both groups were compared. Meanwhile, the effects of Radix Astragali Decoc tion and Injectio on proliferation of vaginal epithelium, PCNA expression, diffe rentiation of tail scale epidermis and plasma ET－1 were assessed in mouse mode l. Results It was shown that psoriatic lesions began to fade significantly earli er in Radix Astragali group than in routine comprehensive treatment group. The c linical cure rate was significantly higher in Radix Astragali group also. Animal experiment indicated that Radix Astragali Injectio had effects on all 4 indices mentioned above and the effects of the Injectio was stronger than that of the D ecoction. Conclusions Radix Astragali Injectio is effective for psoriasis vulgar is. Its therapeutic effects may be explained by blocking multiple pathogenetic l inkage.

Morphea complicated by Hashimoto's thyroiditis is reported in two sisters.Case 1:a 64-year-old female presented with skin rashes on the anterior neck,trunk and bilateral anterior shins for 5 years,itching skin rashes on the perineum for 4 years,and Hashimoto's thyroiditis for 9 years.Physical examination revealed grade 1 enlargement of firm thyroid gland without exophthalmos or pretibial myxedema.Dermatological examination showed pink patches on the neck and breast,sclerosis and atrophy of skin over the back,porcelain-white patches on the perineum.Histopathological findings suggested the diagnosis of morphea on the breast and lichen sclerosus et atrophicus on the perineum.Case 2:a 55-year-old female,who was the younger sister of case 1,suffered from gradual sclerosis and atrophy of skin in the left inframammary region and abdominal region for 4 years,as well as Hashimoto's thyroiditis for 3 years.Similarly,physical examination revealed grade 1 enlargement of firm thyroid gland without exophthalmos or pretibial myxedema.Hypopigmentation,sclerosis and atrophy of skin were observed in the left inframammary region,abdominal region and central back region.Histopathological examination suggested a diagnosis of morphea.According to the clinical and histopathological manifestations,periodic acid-Schiff staining and thyroid gland function test results,the 2 cases were both diagnosed as morphea complicated by Hashimoto's thyroiditis.

Objective To explore molecular mechanisms underlying the in vitro counteracting effect of curcumin on malignant melanoma.Methods Cultured A375 and C8161 human melanoma cells were cultivated in vitro,and randomly divided into several test groups and a control group to be treated with different concentrations of curcumin and dimethyl sulfoxide respectively for different durations.Then,methyl thiazolyl tetrazolium (MTT) assay,Transwell assay,flow cytometry and Western blot were performed to evaluate the effect of curcumin on the proliferation,invasion and cell cycle of,as well as expressions of AKT/mTOR signaling pathway-related proteins in A375 and C8161 cells respectively.Statistical analysis was carried out by using t test.Results MTT assay showed that the treatment with curcumin of 5-35 mg/L for 24-96 hours significantly inhibited the proliferation of both A375 and C8161 cells compared with that with dimethyl sulfoxide (all P ＜ 0.001),and the inhibitory effect was in a dose-dependent manner within the range of 5-15 mg/L for A375 cells and within the range of 5-10 mg/L for C8161 cells,and in a time-dependent manner from 0 to 48 hours for both cells.After treatment for 24 hours,the 50％ inhibitory concentration (IC50) of curcumin against A375 cells and C 8161 cells was 10 mg/L and 5 mg/L respectively.Transwell assay demonstrated that the invasion of A375 and C8161 cells was significantly suppressed by 72-hour treatment with curcumin at 10 mg/L and 5 mg/L respectively (both P ＜ 0.001).Flow cytometry showed that the cell cycle of A375 and C8161 cells was arrested at G2/M phase after 24-hour treatment with curcumin at 10 mg/L and 5 mg/L respectively,with significant differences in the proportion of A375 cells and C8161 cells in G2/M phase between the test group and control group (A375 cells:35.00％ ± 3.54％ vs.120.80％ ± 7.46％,P＜ 0.001;C8161 cells:19.33％ ± 4.04％ vs.85.00％ ± 9.53％,P ＜ 0.001).Western blot revealed that the expressions of AKT/mTOR signaling pathway-related proteins were decreased in A375 and C8161 cells after 24-hour treatment with 10 mg/L and 5 mg/L curcumin respectively.Conclusion Curcumin can inhibit the proliferation and invasion of A375 and C8161 cells,likely by blocking cell cycle and inhibiting activation of the AKT/mTOR signaling pathway.

Objective Mongolian gerbil can make themself urine concentration for saving water and adapt to the harsh desert environment, due to their very unique moisture control system in the body.Methods Mongolian gerbil is resistant to drought on account of their special kidney. Histology of the kidney, ureter and bladder in Meriones Unguiculataus were observed by light microscopy using HE staining.Results The results showed that compared with rats and mice, the Mongolian gerbils have more developed distal tubules, and well developed inner renal medulla.Conclusions We hope that the findings of this study enrich our understanding of the histology of urinary system in Mongolian gerbils and provide support for the laboratory animalization of this animal.

In order to check out the cortical EEG signals of conscious and/or anesthetized rats, a 16-channel analog signals preprocessor is developed. This preprocessor can amplify and filter the primal EEG signals to transform them into ones with proper altitudes and high SNRs, and then they will be sent to the A.D converter. Some measures are taken in view of the signals' low frequencies, low SNRs and background with strong interference. When high gain made, such performances of the circuit are emphasized as low noise, low offset voltage, low gain error and heightened CMRR.

Objective To analyze the results of detection of infectious indicators of patients from the first affiliated hospital of Nanjing Medical University in recent 5 years ,and to provide a scientific basis for the control and prevention of infectious diseases . Methods The patients with clinical data from January 2010 to April 2014 were retrospectively analyzed ,then the infectious indica‐tors of all the subjects were detected and analyzed .Results HIV positive rate was between 0 .043% to 0 .061% ,positive rate of HCV was between 1 .07 to 1 .41% ,positive rate of TP was between 2 .01% to 2 .17% .HBsAg positive rate in 2010 was 8 .36% ,the positive rate was 7 .81% in 2014 .HBsAb positive rate in 2010 was 35 .36% ,positive rate was 50 .96% in 2014 .Conclusion Effec‐tively cut off the route of transmission could prevent the further spread of infectious disease .

Objective To establish a new traceability pathway of point-of-care testing ( POCT ) of HBA1c by using commutable quality controls, in order to improve the accuracy of POCT HBA1c and the harmonization of testing results with those of central laboratories.Methods The study was about measurement traceability. Human frozen whole blood samples with IFCC assigned values were used to calibrate the G8 HBA1c Variant in June, 2013.According to the CLSI EP9-A2-IR guideline, 50 patient samples and 2-level commercial QC samples were then analyzed by G8 system and DCA Ventage system.The best fitting curves for fresh patient samples and the commercial QC materials were established separately. The patient results tested on the DCA Ventage were modified and verified.Paired t-test and Passing Bablok linear regression were used.Results The linear equation of DCA/G8 before calibration was Y=0.899 5X+0.389 1(R2 =0.991 0).Calibration by fresh patient samples reduced the mean bias of DCA/G8 from -0.40%±0.34%to 0.00%±0.29%.Calibration by QCs reduced the mean bias to 0.15%±0.29%.The linear correlation established by quality controls was stable, which made the bias was lower between DCA and G8 in the consequent six runs.Conclusions The accuracy and the traceability of POC testing could be realized by using commutable QC materials traceable to IFCC assigned values.Through this method, POC testing can become more comparable to the results of clinical laboratory HBA1c instruments.

Objective To investigate the roles of glutamate signaling pathway in melanin transfer.MethodsEpidermal melanocytes and keratinocytes were isolated from human foreskin tissue followed by purification and primary culture.Immunofluorescence microscopy was conducted to observe the intracellular distribution of N-methy-D-aspartate receptor 1 (NMDAR1) and NMDAR2A in melanocytes.Some melanocytes were classified into 4 groups to be pretreated with MK801 (the NMDAR antagonist dizocilpine maleate) at 100μmol/L for 5 minutes followed by treatment with NMDA(an NMDAR agonist) at 100 μmol/L (MK801-pretreated group 1),pretreated with MK801 at 100 μmol/L for 1 hour followed by treatment with NMDA at 100μmol/L (MK801-pretreated group 2),treated with MK801 at 100 μmol/L for 5 minutes (MK801 group),treated with NMDA at 100 μmol/L for 5 minutes (NMDA group),respectively,then,confocal microscopy was performed to measure the intracellular calcium (Ca2+) concentration of the melanocytes.The distribution of β-tubulin was visualized by confocal microscopy in melanocytes treated with MK801 at 100 μmol/L for 24 hours.Some melanocytes and keratinocytes were cocultured with or without MK801 at 100 μmol/L for 24 or 48 hours,then,scaning microscopy was carried out to observe the junction structure between melanocytes and keratinocytes,and alkali method coupled with spectrophotometric analysis to determine melanin content in keratinocytes.Results The intracellular calcium concentration of melanocytes was decreased by MK-801,but increased by NMDA at 100 μmol/L,and the increase was blocked by the pretreatment with MK-801 for 5 minutes or 1 hour.After incubation with MK-801 at 100 μmol/L for 24 hours,a more intense staining for β-tubulin was observed around the nuclei of melanocytes.There was a significant reduction in the number of filopodia on the surface of and between melanocytes and keratinocytes after treatment with MK-801 at 100 μmol/L for 48 hours.Also,the content of melanin(represented as the absorbance value at 375 nm) transferred from melanocytes into keratinocytes was statistically reduced in coculture system treated with MK-801 at 100 μmol/L compared with that without treatment (0.158 ± 0.003 vs.2.203 ± 0.006,t =6.323,P ＜ 0.01 ).Conclusions The glutamate signaling pathway exerts a regulatory effect on intracellular calcium concentration of distribution of β-tubulin in,filopodia formation of melanocytes and melanin transfer between melanocytes and keratinocytes.

Objective To investigate the roles of glutamate signaling pathway in the activation of human peripheral blood lymphocytes(PBLs) and pathogenesis of vitiligo.Methods Peripheral blood lymphocytes (PBLs) isolated from 5 patients with generalized vitiligo and 5 healthy controls were cultured in vitro.Flow cytometry was performed to quantify the expression of CD25 and interferon-γ on PBLs derived from healthy controls and treated with MK801 (a non-competitive antagonist of N-methyl-D-aspartic acid receptor,NMDAR) at 100 μmol/L or phosphate buffered saline (PBS) for 48 hours,as well as the level of reactive oxygen species (ROS) in the controlderived PBLs treated with MK801 at 100 μmol/L,NMDA (an agonist of N-methyl-D-aspartic acid receptor) at 0.5 mmol/L or PBS for 48 hours.The protein and mRNA expressions of NMDAR1 and NMDAR2A were measured by flow cytometry and real-time PCR,respectively,in PBLs from the healthy controls and vitiligo patients.Immunohistochemistry was used to observe the expressions of NMDAR1 and NMDAR2A in tissue specimens from depigmented and postinflammatory hyperpigmentation lesions of the patients with vitiligo and from normal skin of the healthy controls.Results Compared with the PBS-treated PBLs from the healthy controls,the MK801-treated PBLs showed a downregulated expression of CD25 (7.28％ ± 0.18％ vs.16.02％ ± 0.42％,P ＜ 0.01),but an upregulated proportion of CD25+IFN-γ+ lymphocytes (1.79％ ± 0.09％ vs.0.78％ ± 0.06％,P ＜ 0.01),and the NMDA-treated PBLs displayed a higher ROS level (101.1 ± 3.50 vs.69.80 ± 2.08,P＜ 0.01 ).The protein expression of NMDAR1 in PBLs was significantly higher in vitiligo patients than in the healthy controls (3.85 ± 2.17 vs.0.97 ±0.55,P ＜ 0.05).Conclusion Glutamate signaling pathway may be involved in the immunopathogenesis of vitiligo via affecting the secretion of interferon-γ by,and ROS level in,activated lymphocytes.