Objective To investigate the effects of S100B on the expressions of dopamine receptors and the synthesis,metabolism of neurotransmitters dopamine,5-hydroxytryptamine which are related to the abnormal motor coordination of Parkinson's disease (PD).Methods The hS100B transgenic mice were established.The mice were divided into S100B transgenic group (TG,n =14),S100B knockout group (KG,n =14) and the non-transgenic control group (CG,n =14).The motor coordination ability of mice was measured by the Rota-rod test.The expressions of dopamine D1 receptor (D1DR),dopamine D2 receptor (D2DR),tyrosine hydroxylase (TH) and phosphorylated TH at Ser19,Ser31,Ser40 in brain tissue were detected by reverse transcription polymerase chain reaction and Western blotting.The levels of Tyr,levodopa,dopamine,homovanillic acid,Trp,5-hydroxytryptamine and 5-hydroxyindoleacetic acid in mesencephalon were measured by high performance liquid chromatography with fluorescence detection.Results Compared with CG,the motor coordination ability of mice (s) showed progressive decline in TG (3 months:4.60±0.30vs4.25±0.21,q =5.194;6 months:4.52±0.31 vs4.07±0.22,q =6.139;9 months:4.43 ± 0.25 vs 3.60 ± 0.18,q =13.484;all P ＜ 0.05),the expressions of D2DR mRNA and protein decreased (1.34 ± 0.13 vs 0.48 ± 0.07,q =21.578;1.05 ± 0.15 vs 0.69 ± 0.10,q =8.063,both P＜0.05) and phosphorylated TH at Serl9 and Ser40 increased (0.95 ±0.10 vs 1.14-0.13,q =4.972;0.94 ± 0.12 vs 1.17 ± 0.14,q=5.382,both P＜ 0.05),the levels of levodopa,dopamine and homovanillic acid were elevated (87.04 ± 11.77 vs 115.28 ± 16.80,q =4.764;56.66 ± 9.87 vs 72.96 ± 11.02,q=3.923;26.58 ± 8.11 vs 38.65 ± 6.67,q=3.981,all P＜ 0.05),the leve1 of 5-hydroxytryptamine was reduced (925.50 ± 74.26 vs 637.87 ± 56.76,q =11.084,P ＜ 0.05),the ratios of homovanillic acid/dopamine and 5-hydroxyindoleacetic acid/5-hydroxytryptamine increased (0.45 ± 0.05 vs 0.54±0.08,q =3.325;0.94±0.07 vs 1.42±0.12,q =12.367,both P＜0.05) in the brain of TG at the age of 9 months old.There was no significant difference of detection indexes between KG and CG.Conclusions S100B plays an important role in the development of PD and the brain-specific S100B transgenic mice can be used to investigate the function of S100B gene on the development of PD.

ABSTRACT:Objective To construct V-set and immunoglobulin domain containing 4 (Vsig4)nanobodies (Nbs) as specific macrophage probes so as to use them as molecular probes of macrophagocytes.Methods A nanobody phage library was generated by using peripheral blood lymphocytes isolated from an alpaca immunized with recombinant Vsig4 protein.After three rounds of selection against recombinant Vsig4.The Nbs were subjected to sequencing and genome alignment to obtain VHH sequence.Nbs were isolated and tested for Vsig4 specificity in an ELISA using recombinant Vsig4.The affinity capacity of Nbs was verified by the cell line stably expressing Vsig4. Results A nanobody phage library with an estimated 7.27 × 107 clones with 70% insertion was successfully constructed.Totally 1 3 6 Vsig4-positive clones were sequenced and aligned according to different CDR3 sequences. In summary,1 5 Vsig4 nanobodies were obtained and grouped into 3 different CDR3 epitopes.The affinity of representing nanobody and Vsig4 was analyzed via ELISA;Nb1 1 9 showed the highest affinity against both recombinant and native Vsig4.Conclusion We successfully constructed and screened Vsig4 specific nanobody number 1 1 9 with high affinity and specificity.It can help with macrophage detection and in vivo monitoring.

BACKGROUND: Our previous studies have demonstrated that cryopreserved bone marrow stromal cells (BMSCs) still maintain high survival rate, cell proliferation and osteogenic differentiation potentials after thawing. However, this result needs confirmed in vivo environment. OBJECTIVE: To explore the effects of cryopreserved BMSCs and collagenic membrane BME-10X complex on type Ⅰ collagen synthesis in vivo. METHODS: Beagle dog BMSCs were cultured in vitro and cryopreserved for 12 months, which were thawed and prepared complexes with collagenic membrane. The complexes were cultured with mineralization induction medium or normal medium for 5 days, followed by implanting into nude mice. The specimens were harvested and analyzed by gross observation, histopathological and immunohistochemistry at 4 weeks after implantation. The collagenic membrane cultured with mineralization induction medium served as controls. RESULTS AND CONCLUSION: In the control group, the boundary of collagenic membrane was distinctly, without cell growth around boundary or intra collagenic membrane, additionally, there was little type Ⅰ collagen. In the non-induction group, cells grew into collagenic membrane, trabes-like collagen formed, and type Ⅰ collagen distribution increased at 4 weeks. In the induction group, scaffold degraded, more cells grew, and plenty of collagen formed osteoid-like tissues. The distribution of typeⅠcollagen was obviously increased than that of other groups. The findings demonstrated that cryopreserved BMSCs possess strong osteogenic differentiation potentials after proliferation and induction combined with collagenic membranes in vitro.

Objective To determine long-term survival of 214 patients of lung cancer with brain metastases and to detect the potential prognostic factors.Methods A retrospective review was pedormed evaluating patients diagnosed as lung cancer with brain metastasis from Jan 1992 to Dec 2001 at Zhejiang Cancer Hospital.Two hundred and fourteen cases were enrolled.All hospital records were thoroughly reviewed in a retrospective manner.The management of the brain metastases were as follows: 8 patients underwent surgical resection and postoperative whole brain radiotherapy (WBRT); 2 cases received resection and chemotherapy; 10 had resection alone; 10 underwent WBRT alone,36 had chemotherapy alone; 15 received the combination of resection,chemotherapy and WBRT; 104 were performed with chemotherapy combined with WBRT; 29 had only supportive care.Survival time was measured from the date of the first treatment for malignancy to the date of death or the last follow-up.Seven further potential prognostic factors were investigated for survival including age,gender,T or N status,number of extra cranial metastases,pathological type and treatment modality.Statistical analysis was performed using the Kaplan-Meier method and Cox-regression analysis.Results The overall median survival time was 10 months (95% CI9.06--10.94) and the 1,3,5 year survival rates were 7.46%,1.14% and 0,respectively.In the univariate model,none of the following variables had effect on survival: age,gender,T stage of the tumor,nodal status,number of extra cranial metastases and histological type.Univariate analysis showed a better survival for the combination of surgical resection,chemotherapy and radiation (P=0.00).Based on Cox-regression analysis,treatment modality was the only independent predictor of survival Conclusions Aggressive combined therapy of brain metastases may achieve a survival advantage.Excellent overall survival of lung cancer with brain metastases has been achieved with a combination of WBRT with surgical resection and chemotherapy.

Objective To study the clinical, histopathologic and ultrastructural characteristics of achromic naevus (AN). Methods Clinical data, including sex, age, age of onset, pattern of lesions, involved sites, shape and number of lesions and associated systemic diseases, were collected from 85 patients with AN. Skin melanin index was detected in 34 lesions of 19 patients with AN, 30 lesions of 12 patients with vitiligo and 64 contralateral normal skin islands of the 31 patients. Reflectance confocal microscopy (RCM) was performed to analyze the lesion, normal skin and junctional area between lesional and normal skin of 62 patients with AN. Tissue samples were obtained from lesions and perilesional normal skin of 17 patients with AN and subjected to pathological examination as well as ultrastructural study with transmission electron microscopy; also, skin biopsy specimens were immunostained for tyrosinase, HMB45, tyrosinase-related protein-1 (TRP-1), TRP-2 and CD117. Results Of the 85 patients with AN, 23 (27.1%) developed lesions at birth, and 21 (24.7%) after 3 years of age; 72 (84.7%) had irregularly shaped lesions, 54 (63.5%) had only a single lesion. The mean melanin index and relative melanin index of AN lesions were 186.56 ± 52.86 and 80 ± 11, respectively, significantly lower than those in normal skin islands (223.88 ± 63.19 and 100, both P < 0.01), but higher than those in depigmented lesions from 12 patients with vitiligo (128.57 ± 64.31 and 60 ± 20, both P < 0.01). RCM revealed a decline in the number of melanocytes and brightness of melanin caps, even distribution of melanin in lesions, as well as obscure demarcation between lesions and normal skin from patients with AN. Fontana-Masson stain showed that the melanin content was lower in lesions than in perilesional skin (1810.12 ± 327.96 vs 2064.24 ± 260.41) from patients with AN. Microscopic examination demonstrated a decrease in melanocyte and melanosome number, presence of immature melanocytes at stage Ⅱ and Ⅲ in cytoplasm and dendrites of melanocytes and keratinocytes, aggregated melanosomes in affected keratinocytes in lesions of AN. In 17 patients with AN, the relative expression levels of tyrosinase and TRP-1 were 1827.35 ± 307.09 and 6102.54 ± 1642.64, respectively, in normal skin specimens, significantly higher than those in lesional skin (1477.35 ± 224.05, 5322.33 ± 1565.26, both P< 0.01); no statistical difference was observed in the expression levels of HMB45, TRP-2 or CD117 between lesional and normal skin. Conclusions AN is an early-onset, nonfamilial aggregated, stable leukoderma with irregular margins, and in lesions of AN, the number of both melanocytes and melanosomes is decreased with the presence of immature melanosomes. The measurement of relative melanin index and reflectance confocal microscopy may offer a non-invasive approach to the diagnosis of AN.

Adolescent ; Adult ; Amino Acid Sequence ; Genotype ; Homozygote ; Humans ; Hypoprothrombinemias ; etiology ; genetics ; Male ; Middle Aged ; Molecular Sequence Data ; Mutation ; Pedigree ; Phenotype ; Young Adult

Objective: To observe the effect of oxymatrine ointment on chronic eczema in mice, and preliminarily explore the mechanism. Methods:Thirty-two Kunming mice were randomly divided into four groups including oxymatrine ointment group, blank model group, blank ointment group and positive drug group treated with compound dexamethasone acetate cream. Eczema skin was con-tinuously treated with drugs for 14 days. The mice were sacrificed on the 15th day, and the eczema skin was clipped from back to make histological sections. The changes of inflammation were observed, and the inflammatory cell count was obtained. Meanwhile, heart blood was collected, and serum was obtained by centrifugation. The serum levels of IL-4 and IL-1β were determined by ELISA. Re-sults:The inflammatory cell count in oxymatrine ointment group and the positive drug group was lower than that in the blank model group and the blank ointment group (P<0. 05). The pathology results showed that oxymatrine ointment could improve chronic eczema symptoms, reduce the inflammatory cell infiltration and decrease edema, which was similar to the effects of compound dexamethasone acetate cream. In the respect of molecular immunology, the IL-4 and IL-1βlevels in serum showed no significant changes in oxymatrine ointment group (P>0. 05). Conclusion: Oxymatrine ointment has certain anti-inflammatory effect on eczema, and the mechanism needs to be studied further.

Objective The study aimed to evaluate the application effect of clinical pathways in laparoscopic cholecystectomy patients by using the meta-analysis.Methods Published randomized controlled trials (RCT) about laparoscopic cholecystectomy patients were searched and screened in China National Knowledge Infrastructure (CNKI),China Scientific Journal Database by VIP,Wanfang Database under present standards.The quality of the included studies was evaluated by certain standards.The Review Manager 5.2 software was used for analysis.Results Totally 29 studies including 5 570 cases were eligible to the criteria (2 753 in the experimental group and 2 853 in the control group) altogether.The meta-analysis showed that the hospitalization time and hospitalization costs in the clinical nursing pathway group were significantly less than those of the control group (SMD=-1.69 and-3.75),the satisfaction degree and the mastering of health knowledge in the clinical nursing pathway group were significantly higher than those of the control group (RR=1.16 and 1.26),the differece had statistical significance.Conclusions Application effect of clinical nursing pathways is superior to the traditional nursing method in laparoscopic cholecystectomy patients.

BackgroundRetinitis pigmentosa (RP) has the genetic and phenotype heterogeneity.To determine the disease-causing gene is a foundation of gene therapy.Objective This study was to localize the pathogenic gene and screen the gene mutation associated with Han Nationality autosomal dominant retinitis pigmentosa (ADRP) in a Chinese family.MethodsTwenty-one families enrolled this study,including 12 patients with ADRP and 9 individuals with normal phenotype.Perimetry,fundus examination,electrooculogram ( EOG ) and electroretinogram (ERG) were performed in 12 patients.Genetic linkage analysis was performed on the subjects in all known genetic loci related to ADRP with a panel of microsatellite markers.Subsequently,the mutation screening of rhodopsin gene was screened by direct DNA sequencing.This study was approved by Ethic Committee of Zhongnan Hospital of Wuhan University.Informed consent was obtained from each subject.ResultsThe fundus appearance of the proband was in accordance with the ADRP,and the EOG and ERG showed undetectable.Contractive visual field also was exhibited in the proband.Linkage analysis showed that the maximum logarithm of the odds(LOD) score reached 3.6671 at marker D3S1292 at recombination fraction θ =0.0.The results of direct DNA sequencing revealed a C→ G transversion mutation at codon 53 in exon 1 of rhodopsin gene,which resulted in a proline to arginine change (Pro53Arg) in 12 patients.However,no similar mutation was found in the unaffected members of this family.ConclusionsThe missence mutation Pro53Arg in rhodopsin gene cosegregate with the RP disease.It is determined to be a pathogenic factor of this ADRP family.

Animals ; Carcinogenesis ; Cell Cycle Proteins ; genetics ; metabolism ; Cyclin E ; metabolism ; F-Box Proteins ; genetics ; metabolism ; F-Box-WD Repeat-Containing Protein 7 ; Gene Silencing ; Genes, Tumor Suppressor ; Humans ; Mutation ; Myeloid Cell Leukemia Sequence 1 Protein ; metabolism ; Neoplasms ; genetics ; metabolism ; pathology ; Proto-Oncogene Proteins c-myc ; metabolism ; Receptors, Notch ; metabolism ; Signal Transduction ; TOR Serine-Threonine Kinases ; metabolism ; Ubiquitin-Protein Ligases ; genetics ; metabolism