Objective To construct a cDNA substractive library of dermal papilla cell (DPC) in patchy and normal area of alopecia areata with technique called suppression substractive hybridization. Methods Total mRNA was isolated form DPC of patchy and normal area. Moreover, double strand cDNAs were synthesized in turn. cDNA from patch and normal area then were restricted and divided into two groups and ligated to the specific adaptor 1 and adaptor 2 respectively. After cDNAs from patchy and normal area hybridized with each other twice and underwent two times of nested PCR, then PCR products were ligated with arms of T/A plasmid vectors to set up the substractive library. Results cDNA substractive library of DPC in patchy and normal area of alopecia areata was set up successfully with highly subtractive efficiency. The amplified library contained 120 positive clones, in which 90 positive subclones contained 100 500bp inserts. Conclusion The cDNA substractive library of DPC in patchy and normal area of alopecia areata is a highly efficient one and lays a solid foundation for screening and cloning new and specific genes from patchy and normal DPC of anagen hair follicle in DPC of alopecia areata.

AIM: To purify the water-extract of Sinisa recipe with macoroporous resigns. METHODS: Screening the purification condition by means of the content of active principle, fingerprint chromatography, the yield of dry extracts,et al. of the purified product. RESULTS: The purified product by type HP20 macoroporous resign is in good agreement with the active fraction of Sinisa in the composition of specific principles. CONCLUSION: HP20 type macoroporos resign is considered as optimum for active fraction of Sinisa recipe in purifying efficience.

Objectives To construct a cDNA subtractive library of dermal papilla cells (DPCs) in anagen with suppression subtractive hybridization (SSH) and clone differentially expressed genes related to DPCs in anagen. Methods Total RNA was isolated from DPC of anagen and telogen follicles. Then ds cDNAs were synthesized in turn using SMART cDNA synthesis technique. After cDNAs from anagen and telogen follicle DPCs were hybridized with each other twice and underwent two rounds of nested PCR, PCR products were ligated with arms of T/A plasmid vectors to set up the subtractive library. Selected clones were verified by reverse Nothern blot and DNA sequencing, and the acquired sequences were analyzed for homology based on Genbank nucleotide database. Results cDNA subtractive library of DPCs in anagen follicle was set up successfully with high subtractive efficiency. Thirty-five genes were identified with 22 known functional genes and 13 unknown functional genes. Conclusions These results demonstrate the effectiveness and sensitivity of SSH in detecting differentially expressed genes from a small amount of clinical samples. Information about such alterations in gene expression might be useful for elucidating the genetic events in hair follicle growth regulation.

Objective To investigate the relations between serum adiponectin and coronary artery calcification score (CACS) and to find the risk factors for coronary artery calcification in maintenance hemodialysis(MHD) patients.Methods Twenty-seven MHD patients(MHD group) and 13 healthy persons (control group) were enrolled in this study.The serum adiponectin was measured by enzyme-linked immunosorbent assay method.CACS was calculated by multi-row spiral CT.The circulating parameters such as hemoglobin (Hb),calcium,phosphate,calcium-phosphate product,intact parathyroid hormone (iPTH),total cholesterol (TC),low density lipoprotein cholesterol (LDL-C),albumin (Alb),high sensitive C-reactive protein (hs-CRP),and so on were detected.Results The level of serum adiponectin in MHD group [(15.00 ± 7.47) mg/L] was significantly higher than that in control group [(2.07 ± 0.83) mg/L],and there was significant difference (P＜ 0.01).Coronary artery calcification(CACS ＞ 0 score) was observed in 88.9％ (24/27) in MHD group and 10/13 in control group.The mean CACS in MHD group was significantly higher than that in control group [655 (0-3 570) scores vs.126 (0-731)scores],and there was significant difference (P ＜ 0.05).Eleven MHD patients and 1 healthy person had severe coronary artery calcifications,(CACS ≥ 400 scores).There was significan t difference in dialysis duration,diastolic pressure,phosphate calcium-phosphate product and adiponectin (P ＜ 0.05 or ＜ 0.01).Spearman analysis showed that CACS of MHD patients was positively correlated with dialysis duration,phosphate,calcium-phosphate product,serum creatinine and adiponectin (P ＜ 0.01 or ＜ 0.05).Only calcium-phosphate product remained as independent predictor of CACS in multivariate analysis (P ＜0.01).Conclusion Coronary artery calcification is common in MHD patients and which is correlated with dialysis duration,serum phosphate,calcium-phosphate product,serum creatinine and adiponectin.

Objective To investigate the location and proliferation of human hair follicle stem cells. Methods The expression of keratin 19(K19) in the human hair follicles on the occiput was detected by immunohistochemical staining, and the positive area was located. The follicle epithelium containing the K19-positive area above the hair bulb were cultured with or without mesenchymes (dermal papilla cells, DPCs) at the air-liquid interface of the collagen gel. When the proliferating colonies formed, the distance from the colony to the bottom of hair follicles was measured and the relationship between the K19-positive position and the proliferation colonies was analyzed. The ultrastructure of the proliferation colony was observed by transmission electron microscopy. Results Two portions were found positive for K19 in the outer root sheath of the human hair follicles, the upper one being at the bulge region and the lower one at the outer root sheath above the hair bulb. The proliferating colony formed only when the epithelium of the hair follicle was co-cultured with mesenchymes cells. The statistical analysis suggested the lower portion positive for K19 and the proliferation colonies in the culture were traced to locate at the same site of the human hair follicles. Under electron microscopy, juvenile cells, mature keratinocytes and apoptotic cells were found in the proliferating colonies. Conclusions The human hair follicles may contain two distinct reservoirs for stem cells, which locate in the bulge area and in the region of the outer root sheath above the hair bulb, respectively. Mesenchymes are needed for the proliferation of stem cells. New stem cells, mature keratinocytes and apoptotic cells may be three endings of the stem cells′ proliferation.

Objective To summarize and analyze the pathological characteristics of solid pseudopapillary tumor of pancreas (SPTs).Methods The clinical data of 51 cases of SPTs were retrospectively analyzed.The immunohistochemical localizations of different markers (HSE,SYN,CD_(56),CD_(10),Nestin,Vim,a1-ACT,EMA,AE1/AE3 and CK19) on 39 SPTs were studied.Results Pathological features included a combination of solid and cystic components with pseudopapillae formation and degenerative regions without glands.Among the 39 cases of SPTs,the expression rate of NSE was 97.4%,the expression rate of CD_(56),CD_(10) was 84.6%,the expression rate of Nestin and Vim was 64% and 87%,the expression rate of S100 was 79.5%,the expression rate of a1-ACT and a1-AT was 82.1% and 79.5%,while the expression rate of SYN was 12.8%;however there was low expression and weak positive reaction of EMA,AE1/AE3 and CK19.Conclusions The typical pathological characteristics of SPTs may result from gradual degenerative changes induced anoxemia in some SPT's areas.The heterogeneity of SPTs on different antibody markers showed that the SPTs may be originated from pancreatic embryonic stem cells,and result from immature differentiation of the pluripotential stem cells during pancreatic genesis.

AIM: To investigate the clinical characteristics of patients with systemic lupus erythematosus ( SLE) complicated with retinopathy. METHODS: Totally 121 cases of SLE patients treated in our hospital from February 2015 to February 2017 were selected,including 30 cases with retinopathy(observation group) and 91 cases without retinopathy(control group), the clinical manifestations and laboratory examination results of the two groups were compared. RESULTS: ln the observation group, there were 6 patients with bilateral retinopathy and 24 patients with monocular retinopathy. Cotton retinal exudation, retinal vascular occlusion and retinal hemorrhage lesions were common, accounted for 33%, 25% and 19%. The incidence of skin rash, skin vasculitis and Raynaud's phenomenon in the observation group were 63%, 47% and 37%, respectively, which were significantly higher than those in the control group (P<0.05). There were no significant difference in the incidence of mucosal ulcer, arthritis, neuropsychiatric symptoms and pleurisy between the observation group and the control group(P>0.05). The positive rate of anti ds-DNA in the observation group was 63%, which was significantly higher than that in the control group (P<0.05). The observation group and the control group urine protein > 3 +, anti Sm antibody positive and rRNP positive and antiphospholipid antibody percentage differences were not statistically significant(P>0.05). The SLEDAI score of the observation group was 20.14 (9, 30) points, which was significantly higher than that of the control group (P<0. 05). The modified BenEzra score of the observation group was 10 04士3.15,and was positively correlated with the SLEDAI score (rs=0.706,P<0.05). CONCLUSION: SLE with fundus retinopathy patients mainly manifest as cotton wool spots, retinal vascular occlusion and retinal hemorrhage; rashes, vasculitis, Raynaud's phenomenon,and anti ds-DNA antibodies are common in SLE patients with retinopathy. The modified BenEzra score plays an important role in the evaluation of SLE disease activity and retinal vascular lesions.

OBJECTIVETo study the regularity of migration and distribution of bone marrow stromal cells (BMSCs) in injured spinal cord with intradural space transplantation.

METHODSForty Wistar rats were randomly assigned into 4 groups. The spinal cord injury model was prepared according to the modified Allen method. BMSCs were labeled by CM-Dil. And 5.0 multiply 10(6) cells were transplanted by different channels including intraventricular injection (Group A),injured spinal cord intrathecally injection (Group B), remote intrathecally injection at the L(3)-L(4) level (Group C), and intravenous injection (Group D). Spinal cord was dissected at 24 hours, 1, 2, 3 and 4 weeks after transplantation. Sections of 4 micromolar were cut on a cryostat and observed under fluorescence microscopy.

RESULTSNo fluorescence was observed 24 hours after transplantation in spinal cord injury parenchyma except Group B. One week later, BMSCs in Groups A and C began to migrate to the injured parenchyma; 2-4 weeks later, BMSCs penetrated into the injured parenchyma except Group D. The number of BMSCs decreased at 3-4 weeks after transplantation. The number of cells in Group B decreased faster than that of Groups A and C.

CONCLUSIONSBMSCs transplanted through intraventricular injection, injured spinal cord intrathecally injection and remote intrathecal injection could migrate to the injured parenchyma of spinal cord effectively. The number of BMSCs migrated into injured spinal cord parenchyma is rare by intravenous injection.

Animals ; Bone Marrow Cells ; cytology ; Bone Marrow Transplantation ; methods ; Cell Movement ; physiology ; Male ; Random Allocation ; Rats ; Rats, Wistar ; Spinal Cord Injuries ; pathology ; surgery ; Stromal Cells ; cytology ; transplantation

[Objective] To evaluate the efficacy of autologous whole blood injections in patients with chronic spontaneous urticaria and positive autologous serum skin test (ASST).[[Methods]] After assessment of clinical history,patients with chronic spontaneous urticaria underwent skin prick test (SPT) and ASST.Then,100 patients with positive ASST but negative SPT for common allergens were randomly classified into treatment group (n =60) and control group (n =40).Oral loratadine was given to all the patients with a gradual tapering to the least maintenance dose.Patients in the treatment group were also injected with autologous whole blood once a week for 12 times.Patients were evaluated by urticaria activity score (UAS) and dermatology life quality index (DLQI) at the baseline,the end of the 3rd and 6th month after the initial treatment.The total amount of antihistamines required for the control of urticaria every month was calculated.The UAS,DLQI,accumulative amount of administrated antihistamines,and the diameter of wheal/flush induced by autologous serum were compared by t test before and after the treatment,and the efficacy was compared by rank sum test between the two groups.[Results] No significant difference was observed between the control and treatment group in UAS at the baseline (5.73 ± 0.51 vs.5.32 ± 0.79,P＞ 0.05).The UAS reached 1.57 ± 1.42 and 0.69± 0.92 with a decrease rate of 69％ and 81％ in the treatment group,and 3.65 ± 1.53 and 2.65 ± 1.61 with a decrease rate of 35％ and 53％ in the control group,respectively at the end of the 3rd and 6th month,and statistical difference was observed for the decrease in both groups at the two time points (all P ＜ 0.05).The total amount of antihistamines required for the control of urticaria per month averaged 8.63 pills and 3.83 pills respectively in the treatment group after 3 and 6 months of treatment,significantly less than that in the control group (16.85 and 15.27 pills,respectively).[Conclusion]s The combination of oral antihistamine and autologous whole blood injections can not only reduce disease activity and improve patients' quality of life,but also decrease the total amount of antihistamines required for the control of urticaria.

OBJECTIVETo study the roles of TNF-alpha secretion of PBMC in patients with chronic hepatitis C and Nonalcoholic fatty liver.

METHODSThe level of TNF-alpha secretion of PBMC in patients with chronic hepatitis C and Nonalcoholic fatty liver was detected by ELISA after culturing for 72 hours in vitro, as well as patients with chronic hepatitis C and the normal control.

RESULTS(1) Compared with the normal control, the level of TNF-alpha in group with chronic hepatitis C and in group with chronic hepatitis C and Nonalcoholic fatty liver notably increased. (2) Compared with the group with chronic hepatitis C, the level of TNF-alpha in group with chronic hepatitis C and Nonalcoholic fatty liver also notably increased.

CONCLUSIONIt suggested TNF-alpha takes important roles in the infection course of chronic hepatitis C with Nonalcoholic fatty liver.

Adolescent ; Adult ; Cells, Cultured ; Child ; Fatty Liver ; metabolism ; Female ; Hepatitis C, Chronic ; metabolism ; Humans ; Leukocytes, Mononuclear ; metabolism ; Male ; Middle Aged ; Tumor Necrosis Factor-alpha ; metabolism ; Young Adult