Background Stem cell transplantation represents a promising treatment option for patients suffering from degenerative disorders.Accumulating evidences indicate that mesenchymal stem cells (MSCs) are able to differentiate into retinal pigment epithelial (RPE)-like cells.However,MSCs are difficult to obtain.Human adipose mesenchymal stem cells (ADSCs) are proved to have similar properties to MSCs,but relevant study is less.Objective This study was to assess the feasibility of human ADSCs differentiating into RPE-like cells and the safety of its application in vivo.Methods The third generation of human ADSCs were incubated into 6-well plate,and 100 ng/ml epithelial growth factor,50 μ mol/L taurine and 5×10-7 mol/L retinoic acid were added into the medium 12 hours after cultured to induce the cells,and conventional cultured cells were used as the control group.Induced cells were traced with PKH26,and Pan-cytoke ratin (Pan-CK) monoclonal antibody was used to identify the cells under the fluorescence microscope.Induced RPE-like cell suspension of 1 μl was intravetreally injected in the right eyes of 6 BALB/c mice,and equal volume of PBS was used in the same way in another 6 mice.The animals were sacrificed 1 month after injection,and the retinal morphology was examined by histopathology under the optical microscope.The ultrastructure of retinal ganglion cells (RGCs) was examined by the transmission electron microscope.The use and care of the animals complied with Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission.Results Cultured human ADSCs grew well with the slender polygone shape.Cell membranes showed the red fluorescence for PKH26 after induced.In addition,Pan-CK was expressed in the cell membranes with the red fluorescence in the induced cells,but the response was absent in the control cells.One month after intravitreal injection,induced cells located on the retinal surface,and the retinal morphology was clear under the optical microscope.No abnormality in RGCs was seen under the transmission electron microscope.Conclusions Human ADSCs can differentiate into RPE-like cells after induction.PKH26 can mark induced cells well.There is no adverse effect of induced cells on retina after intravitreal injection in a short-term duration in mice.

Objective: To observe the clinical efficacy of long-time needle retaining at Baihui (GV 20) in treating post-stroke cognitive impairment. Methods: A total of 60 eligible patients with post-stroke cognitive impairment were randomized into a treatment group and a control group by random digital table, 30 cases in each group. The control group was intervened by basic treatment including routine therapy in Neurology department and Western medicine rehabilitation therapy, while the treatment group was intervened by same treatment in the control group combined with long-time needle retaining at Baihui (GV 20). Mental state of patients was evaluated by National Institute of Health stroke scale (NIHSS) and mini-mental state examination (MMSE) after 4 weeks of treatment. Results: After 4-week treatment, the NIHSS score dropped in both groups, and intra-group comparisons showed statistical significances (both P<0.05), and the score in the treatment group was statistically lower than that in the control group (P<0.01); the MMSE score in the treatment group was significantly higher than that in the control group, there was a statistical inter-group difference (P<0.01); the total effective rate was 93.3%, and the marked effective rate was 50.0% in the treatment group, versus 76.7% and 20.0% in the control group, there were significant differences in the total effective rate and the marked effective rate between the two groups (all P<0.05). Conclusion: Long-time needle retaining at Baihui (GV 20) can produce a safe valid therapeutic efficacy in treating post-stroke cognitive impairment.

Objective:To observe the clinical efficacy of warm needling therapy for chronic lumbar strain.Methods:A total of 60 patients with chronic lumbar strain who met the inclusion criteria were randomized into a treatment group and a control group by the random number table,with 30 cases in each group.The treatment group was treated with warm needling therapy,while the control group was treated with ordinary acupuncture treatment.The treatments were both performed once every other day,and 7 times constituted a course of treatment.Visual analog scale (VAS) score was used to assess the degree of pain and the clinical efficacy was compared between the two groups.Results:The total effective rate of the treatment group was higher than that of the control group (P＜0.05).There was no significant difference in VAS score between the two groups before treatment (P＞0.05).After treatment,the VAS scores of both groups decreased significantly,and the intra-group differences were statistically significant (both P＜0.05).The VAS score of the treatment group after treatment was statistically different from that in the control group (P＜0.05).Conclusion:Warm needling therapy has a better curative effect than ordinary acupuncture in the treatment of chronic lumbar strain.

Objective To evaluate the efficacy of extended-field intensity-modulated radiotherapy (IMRT) in the treatment of patients with early-stage NK/T cell lymphoma (NKTCL),and to examine the clinical characteristics and the effect of treatment factors on the prognosis of these patients.Methods The clinical data of 165 patients with early-stage NKTCL who underwent extended-field IMRT with (n=158,95.8%) or without chemotherapy (n=7,4.2%) were reviewed.Of these 165 patients,140(84.8%) received a radiation dose of ≥50 Gy to the primary lesion,and 25 patients (15.2%) received a radiation dose of<50 Gy.Most patients (n=147,89.1%) were treated with L-asparaginase-based chemotherapy regimens,whereas only 11 patients (6.7%) were treated with doxorubicin-based CHOP/CHOP-like regimens.In addition,109 patients (66.1%) received ≥4 cycles of chemotherapy.Locoregional control (LRC),overall survival (OS),and progression-free survival (PFS) rates were calculated using the Kaplan-Meier method,and the log-rank test was used for survival comparison and univariate prognostic analysis.A multivariate prognostic analysis was performed using the Cox model.Results The 5-year sample size 55.The 5-year OS,PFS,and LRC rates of all patients were 74.2%,72.5%,84.4%,respectively.The patients who received a dose of ≥50 Gy had a significantly higher 5-year LRC rate than those with<50 Gy (91.8% vs.39.7%,P=0.000).The 5-year OS was significantly higher in the low-risk early-stage group than in the high-risk early-stage group (P=0.002).For the high-risk early-stage NKTCL group,patients who received ≥4 cycles of chemotherapy had significantly higher 5-year OS and PFS than those who received<4 cycles of chemotherapy (5-year OS:71.3% vs.59.5%,P=0.032;5-year PFS:70.4% vs.54.4%,P=0.009).In addition,multivariate analysis showed that ECOG≥2,primary tumor invasion (PTI),and Ann Arbor stage Ⅱ were associated with poor OS (P=0.006,0.002,0.014),and ECOG≥2 and PTI were associated with reduced LRC (P=0.004,0.016).Furthermore,ECOG≥2,PTI,Ann Arbor stage Ⅱ,and extranasal primary site were associated with lower PFS (P=0.045,0.003,0.030,0.032).Conclusions Extended-field IMRT at a dose of ≥50 Gy can lead to favorable LRC,OS,and PFS in patients with early-stage NKTCL.However,it is less effective against distant early-stage NKTCL in patients with poor prognosis.Nevertheless,≥4 cycles of chemotherapy can significantly improve the OS and PFS of patients with early-stage NKTCL.

The clinical application of orbital magnetic resonance (MR) T2-mapping imaging in detecting the disease activity of Graves' ophthalmopathy (GO),and the predictive values of therapy response to intravenous glucocorticoid (ivGC) were investigated.Approved by the local institutional review board (IRB),106 consecutive patients with GO were included in this prospective study.All subjects were divided into two groups according to the patients' clinical activity score (CAS):the CAS positive group (CAS ≥3) or the CAS negative group (CAS ＜3).T2 relaxation time of extraocular muscles (T2RT;ms) and the areas of four extra-ocular muscles (AEOMs;mm2) were measured by 3D T2-mapping MR sequence before and after methylprednisolone treatment,so as the CAS and some ophthalmic examinations including visual acuity,intra-ocular pressure,eyeball movement,diplopia and proptosis.In addition,24 healthy volunteers were recruited as the control group.The mean T2RT and AEOMs in CAS positive group were higher than those in CAS negative group.Both CAS positive and negative groups had significantly higher mean T2RT and AEOMs than the control group (P＜0.01).There was a positive correlation between T2RT and AEOMs values in GO patients,both of them had a positive correlation with CAS and the ophthalmic examinations.It was concluded that to evaluate the activity of GO,CAS was mostly related to inflammation symptoms of ocular surface,more than that,T2RT and AEOMs were also related to abnormal findings of the ophthalmic examinations including high ocular pressure,impaired eyeball movement,diplopia and proptosis.T2RT and AEOMs can reflex the inflammation state of ocular muscles better.CAS combined with 3D T2-mapping MR imaging could improve the sensitivity of detection of active GO so as the prediction and evaluation of the response to methylprednisolone treatment.

OBJECTIVETo further study the therapy of wasting muscle by myostatin as a new targets, the eucaryotic expression vector coupled the foreign T-helper epitope of tetanus toxin (TT) to the N terminus of myostatin was constructed, and the effects of the gene vaccine on forelimb grip were tested in immunized mice.

METHODSA DNA fragment encoding the TT epitope followed by the N terminus of mature myostatin (330bp) was synthesized. The eucaryotic expression vector of myostatin was constructed and the chinese hamster ovary (CHO) cells were infected with the recombinant plasmids pVAC-TT-Ms by liposome transfection according to routine laboratory procedure. The myostatin expression was tested by cell immunofluorescence technique in transfected CHO. The forelimbs grip were tested in immunized mice with myostatin gene vaccine.

RESULTSThe eucaryotic expression vector of myostatin coupled TT epitope was constructed successfully through the restriction analysis and sequencing. The recombinant plasmids pVAC-TT-Ms met quality criterion as gene vaccine by analysis OD260/280 and electrophoresis. The myostatin expression was detected obviously in transfected CHO. The forelimb grip in immunized mice had an obvious increase. The average value of forelimb grip of the mice immunized with pVAC-TT-Ms was about 29.88% greater than that of control mice.

CONCLUSIONThe construction of eucaryotic expression vector of myostatin coupled TT epitope is successful in expression for recombinant human mature peptide of myostatin. The gene vaccine of myostatin meet quality criterion. The immunized mice has an obvious increase in forelimb grip.

Animals ; CHO Cells ; Cricetinae ; Cricetulus ; Epitopes, T-Lymphocyte ; Genetic Vectors ; Hand Strength ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Muscle, Skeletal ; physiology ; Myostatin ; genetics ; immunology ; Plasmids ; Transfection ; Vaccines, DNA ; genetics ; immunology

AIMTo establish a simple method for molecular identification of original plants of D. chrysanthum and D. fimbriatum using molecular marker rDNA ITS region.

METHODSRestriction patterns of ITS fragments were obtained using PCR-RFLP method. The PCR products of D. chrysanthum and its morphologically allied species were digested at 37 degrees C by Cla I and Apa LI, those of D. fimbriatum and its morphologically allied species were digested by Sph I.

RESULTSD. chrysanthum, D. fimbriatum and their morphologically allied species could be identified by predicted restriction profiles of PCR-RFLP. The botanical origin of twenty-five fresh samples of "Shihu" collected in markets was identified by this method.

CONCLUSIONThe results showed that PCR-RFLP analysis of the rDNA ITS region is a feasible, simple and inexpensive method for determining the botanical origin of the traditional Chinese medicine "Shihu".

DNA, Plant ; analysis ; DNA, Ribosomal ; analysis ; Dendrobium ; classification ; genetics ; Drug Contamination ; Plants, Medicinal ; classification ; genetics ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Sequence Analysis, DNA ; Species Specificity

OBJECTIVETo define molecular characters to distinguish D. chrysanthum from its allied species D. primulinum, D. lituiflorum, D. aphyllum, D. crepidatum.

METHODThe molecular characteristics of D. chrysanthum and its allied species were compared. The sequences of rDNA ITS regions were exploited to explore the evidence for authentication D. chrysanthum and its allied species.

RESULTAlthough the morphological difference was slight, the sequence difference of ITS regions among five rDNAs was obvious and stable. Fifteen sites of ITS region were defined as DNA character to identify D. chrysanthum from the other four allied species.

CONCLUSIONThe difference of rDNA ITS sequences can be used to authenticate accurately D. chrysanthum from three allied species of Dendrobium.

Base Sequence ; DNA, Plant ; genetics ; Dendrobium ; classification ; genetics ; Drug Contamination ; Molecular Sequence Data ; Plants, Medicinal ; genetics ; Sequence Analysis, DNA ; Species Specificity

OBJECTIVETo study rDNA ITS sequence differences between F type and that of H type of Dendrobium officinale in main habitat of China.

METHODThe population differences of the rDNA ITS region (including ITS1, ITS2, 5.8S) sequences of D. officinale were studied by the method of DNA sequences analysis.

RESULTThere were two different sites between the rDNA ITS sequence of F type and that of H type. One was in ITS1 region, and the other was in 5.8S region. It was proved that there was some relativity between the character of rDNA ITS region and the life type of the populations. The phenomenon of single nucleotide polymorphism (SNP) existed in 5.8S region of rDNA ITS region between F type and H type. The sequences of rDNA ITS region of D. officinale were reported for the first time, and the sequences of ITS region ranged 634 bp (ITS1 231 bp, ITS2 240 bp, 5.8S 163 bp).

CONCLUSIONThe analysis of rDNA ITS of D. officinale deeply reveal the population differences of D. officinale of F type and H type.

Base Sequence ; DNA, Plant ; genetics ; DNA, Ribosomal ; genetics ; Dendrobium ; classification ; genetics ; Molecular Sequence Data ; Plants, Medicinal ; genetics ; Polymorphism, Single Nucleotide ; Sequence Analysis, DNA ; Species Specificity

Background The incidence of retinal vascular diseases increase annually,such as diabetic retinopathy,retinopathy of prematurity and age-related macular degeneration.The key of treatment for these diseases is how to evaluate retinal vascular change effectively and objectively.Retro-orbital injection of fluorescein isothiocyanatedextran (FITC-dextran) is a simple and effective method for observing C57BL/6J mouse retinal vessels.But,whether it is suitable for other mice and rats is seldom reported.Objective This experiment was to assess the feasibility of the observation of retinal vessels by retro-orbital injection of FITC-dextran in different genus of mouse and offer the reference for relevant study.Methods Twelve animals of C57BL/6J mice,Kunming mice,SD rats and Wistar rats were selected,respectively and divided into the experimental group and control group at average.The right eyes of the animals of the experimental group received the retro-orbital injection of 9 ml/kg FITC-dextran,and the right eyes of animals of the control group received PBS solution at the same volume and way.All the animals were sacrificed 10 seconds after injection and both eyes of each animal were obtained for retinal stretched preparation.The retrobulbar tissue and whole-mount retina were viewed under a fluorescence microscope.The use of the animals complied with Regulation for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission.Results Retinal blood vessels labeled by FITC-dextran could be observed in both eyes of C57BL/6J mice and Kunming mice to present with a green fluorescence in experimental group under a fluorescence microscope,but no any fluorescence-labeled retinal blood vessel was exhibited in the control mice.The retinal blood vessel could not be observed in all eyes of SD rats and Wistar rats after the injection of FITC-dextran both in the experimental group and the control group under a fluorescence microscope.The surrounding tissues of the right eyes of mice and rats dyed with green fluorescence of FITC-dextran in the experimental group,however,green fluorescence could not be seen in the surrounding tissues of the left eyes of mice and rats.Conclusions Retro-orbital injection of FITC-dextran is a suitable method of observing the retinal vessels of mouse but not rat.