Objective To develop a visible detection system prepared by protein microarrays which was based on GoldMag particles-labeled technique and compare the results of detection using the protein which was labeled with GoldMag particles and colloidal gold.Methods Human IgG was printed on the glass slides modified with epoxy groups,and goat anti-human IgG conjugated with GoldMag particles and colloidal gold respectlively was then added on the slide.The glass slides were incubated,and then the black images of microarray spots were produced by immuno-gold-silver staining method and observed by naked eyes and recorded with common flatbed scanner.Results The high signal-to-noise ratio could be obtained when the optimized procedures of GoldMag particles-labeling probe were introduced to the protein chip.The conditions of optimum assay were as follows:the spotting concentration of human IgG was 0.2 mg/ml,the glass slides were incubated at 37 ℃ for 2 hour to immobilize human IgG,and the silver enhancement time was 10 min-15 min.Parallelly,the optimized conditions for colloidal gold were as follows:the spotting concentration of human IgG was 0.1 mg/ml,the slides were incubated at 37 ℃ for 1 hour to immobilize human IgG,and the silver enhancement time was 15 min-20 min.Conclusions GoldMag particles-labeled protein technique is comparable to colloidal gold in applying to protein microarrays.The method is considerably simple and practical,and the protein labeled by GoldMag particles could be quantitative.

Objective To establish a diabetic rat model of hindlimb ischemia, and provide a test method for dia-betic limb ischemia and diabetic foot study.Methods Twenty-five Wistar rats were divided into three groups:the model group and control group ( n=10) , and sham operated group ( n=5) .The rats of model group were induced by intraperito-neal injection of streptozotocin ( STZ, 60 mg/kg) , with the blood glucose level over 16.8 mmol/L, while the rats of control group were injected with normal saline.All the rats had the left femoral artery and its branches ligated, and then Doppler scan blood flow analysis was performed for the two hind limbs after operation at different time points.In the meantime, the changes of body weight and blood glucose were observed.The animals were sacrificed at 21days after operation.HE stai-ning was used to observe the pathological changes of gastrocnemius and femoral arteries.Capillary density and intima hyper-plasia were examined using immunostaining for CD31 and a-SMA.Results Blood glucose of the model group rats was sig-nificantly increased as well as the quantity of urine and feces,and associated with weight loss, which were maintained for more than 21 days.The blood flow of control rats was markedly decreased immediatly after operation, reached to the lowest point after surgery, and recovered from 7 -14 days after operation.In contrast to the control group, the model group showed a marked reduction in blood flow in the ischemic hind limb.Pathological examination revealed remarkable vascular atrophy and a significantly reduced number of vessels per high power field in the gastrocnemius muscle of model group rats with respect to the control group, and the femoral arteries of model rats were more narrowed than that of the control rats. Conclusions In this study, an effective, convenient diabetic rat model of hindlimb ischemia is successfully established which will facilitate the studies of drug intervention for diabetic limb ischemia and diabetic foot in the future.

[ ABSTRACT] AIM:To evaluate the effects of 1α, 25-dihydroxyvitamin D3 on T helper cell 17 ( Th17 cells) and its related cytokines in a mouse model of corneal allograft transplantation.METHODS:C57BL/6 mice were transplanted with corneal grafts from BALB/c mice and treated intraperitoneally with 1.0μg 1α, 25-dihydroxyvitamin D3 or soybean oil every other day after operation.The transparency of the corneal grafts was evaluated for potential rejection signs by slit lamp biomicroscopy and histopathology.The expression levels of IL-17, RORγt and IFN-γin the spleen were measured by real-time PCR.Moreover, the protein expression of RORγt and IL-17 in the peripheral blood was analyzed by Western blotting. IL-17 and IFN-γin peripheral blood were measured by flow cytometry.RESULTS:1α, 25-dihydroxyvitamin D3 signifi-cantly inhibited the rejection of the corneal allograft and reduced the numbers of inflammatory infiltrates in the corneal graft. In the spleen, 1α, 25-dihydroxyvitamin D3 treatment reduced the expression levels of IL-17, RORγt and IFN-γ.In the pe-ripheral blood, 1α, 25-dihydroxyvitamin D3 treatment downregulated the expression levels of RORγt, IL-17 and IFN-γ. CONCLUSION:The effects of 1α, 25-dihydroxyvitamin D3 on suppressing corneal transplantation-induced allograft rejec-tion in mice are closely associated with its modulation on IL-17 and related cytokine RORγt.

OBJECTIVEThe SSR information in the transcriptome of Erigeron breviscapus was analyzed in this study, in order to further develop new functional genes SSR markers laid a solid foundation.

METHODSSR loci were searched in all of 52,060 unigenes by using est_timmer. Perl program and SSR primers were designed by Primer3. Furthermore, 36 pairs of primers were randomly selected for the polymorphism analysis on 13 Erigeron breviscapus plants collected from different places.

RESULTA total of 3639 SSRs were found in the transcriptome of Erigeron breviscapus, distributed in 3260 unigenes with the distribution frequency of 6.99%. Di-nucleotide repeat was the main type, account for as much as 34.41% of all SSRs, followed by mono-nucleotide (31.41%) and tri-nucleotide repeat motif (28.08%). The di-nucleotide repeat motifs of AT/AT and AC/GT were the predominant repeat types (28.71%). The tri-nucleotide repeat motifs of AAT/AT was the predominant repeat types (7.94%). For validation the availability of those SSR primers, we randomly selected 36 pairs of primers for PCR amplification. Among them, 34 pair primers (94.44%) produced clear and reproductive bands, 19 pair primers showed polymorphism (52.78%), and 13 Erigeron breviscapus plants were divided into 2 groups.

CONCLUSIONThere are numerous SSRs in Erigeron breviscapus transcriptome with high frequency and various types, this will provide abundant candidate molecular markers for genetic diversity study and genetic map in this plant.

China ; DNA Primers ; genetics ; Erigeron ; classification ; genetics ; Genetic Variation ; Microsatellite Repeats ; Phylogeny ; Polymorphism, Genetic ; Transcriptome

OBJECTIVETo study clinical effects of one-hole microplate internal fixation for the treatment of collateral ligament injuries of the metacarpophalangeal joint of the thumb combined with fracture.

METHODSTwenty-two patients (16 males, 6 females) with collateral ligament injuries of the metacarpophalangeal joint of the thumb combined fracture were treated with one-hole microplate internal fixation. The age of the patients ranged from 18 to 53 years old with a mean age of 28.5 years old. The duration from injury to surgery ranged from 2 hours to 2 months, and the mean time was 6 days. All the patients had collateral ligament injuries combined with fracture of the metacarpophalangeal joint of the thumb. Thirteen patients had injuries in the right hand and 9 patients had injuries in the left hand. There were 18 cases of closed wound and 4 cases of open wound. Eighteen patients had fresh injuries (< 2 weeks) and 4 had old injuries (> 2 weeks). Sixteen patients had injuries in the ulnar collateral ligament of the thumb combined with fracture, 6 patients had radial collateral ligament injuries of the thumb combined with fracture, 4 cases of which were complicated with injuries of abductor pollicis brevis and the end of the flexor pollicis brevis tender. The size of the avulsed fragment was about 3.0 mm x 4.0 mm to 6.0 mm x 7.0 mm.

RESULTSThe incisions of 22 patients healed by first intention. The follow-up periods ranged from 6 months to 5 years old,with an average of 2.5 years old. The thumb function was evaluated by Saetta and other evaluation criteria, and 20 patients got an excellent result and 2 good.

CONCLUSIONThe application of one-hole microplate internal fixation in treating collateral ligament injuries with fracture of the metacarpophalangeal joint of the thumb is an effective method.

Adolescent ; Adult ; Bone Plates ; Collateral Ligaments ; injuries ; surgery ; Female ; Fracture Fixation, Internal ; methods ; Fractures, Bone ; surgery ; Humans ; Male ; Metacarpophalangeal Joint ; injuries ; surgery ; Middle Aged ; Thumb ; injuries ; surgery ; Young Adult

OBJECTIVETo investigate the regulatory role of JAK2/STAT3/vimentin signaling pathway on the proliferation and migration of human colon cancer cells.

METHODSThe human colon cancer cell Lovo was treated with Janus kinase inhibitor AG490. The proliferation of Lovo cells was quantified by MTT assay. The migration of Lovo cells was measured with scratch assay. The intracellular phosphorylation STAT3 (P-STAT3) and vimentin protein was detected by Western blot and immunofluorescence.

RESULTSAfter AG490 treatment, the proliferative ability of Lovo cells decreased as compared with control group (P<0.05). This suppression was dose-independent and time-independent. At 24 hrs after scratching, scratch width in the AG490 group recovered to 20%, lower than that in the control group (60%, P<0.05). After AG490 treatment, the expression of P-STAT3 and vimentin in Lovo cells decreased significantly (P<0.05).

CONCLUSIONJAK2/STAT3/vimentin signaling pathway participates in regulating the proliferation and migration of human colon cancer cells.

Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Colonic Neoplasms ; metabolism ; pathology ; Humans ; Janus Kinase 2 ; metabolism ; STAT3 Transcription Factor ; metabolism ; Signal Transduction ; Vimentin ; metabolism

Conventional sample preparation technique for adipose tissue is difficult to achieve satisfactory results for scanning electron microscopy (SEM). We adopted a strategy of postfix with osmium tetroxide to stabilize the fatty acids, phospholipids protein, and hence the membrane structure. Also by extending the dehydration time to fully replace the organic solvents, we achieved satisfactory results for SEM of adipose samples.
Adipose Tissue ; ultrastructure ; Animals ; Female ; Humans ; Microscopy, Electron, Scanning ; Rats ; Rats, Sprague-Dawley ; Specimen Handling ; methods

OBJECTIVETo evaluate the clinical significance of STAT3, E-cadherin and vimentin in colon cancer.

METHODSSamples of colon cancer tissue and adjacent normal tissue were procured from 70 patients with colon cancer. The expressions of STAT3, E-cadherin and vimentin were detected by immunohistochemistry. Associations of clinicopathological characteristics and these three factors were evaluated.

RESULTSSTAT3, E-cadherin, vimentin were positive in 74.3%,32.9%, and 78.6% in the colon cancer tissues, respectively, and were 15.7%, 82.9%, and 12.9% in normal colon mucosa tissues, respectively. They were correlated with tumor differentiation, depth of invasion, lymph node metastasis, and TNM staging(P<0.05). There was a positive correlation between STAT3 and vimentin expression(P<0.01). However, STAT3 was negatively correlated with E-cadherin(P<0.01).

CONCLUSIONSSTAT3, E-cadherin and vimentin are closely associated with colon cancer. The expressions of STAT3, E-cadherin and vimentin may serve as prognostic indicators for patients with colon cancer.

Adult ; Aged ; Aged, 80 and over ; Cadherins ; metabolism ; Colonic Neoplasms ; metabolism ; pathology ; Female ; Humans ; Male ; Middle Aged ; STAT3 Transcription Factor ; metabolism ; Vimentin ; metabolism

The present study demonstrated that bone marrow cells from diabetic rats were able to form cell clusters expressing insulin,C-peptide,glucagon,somatostatin and islet amyloid .polypeptide,and other genes associated with development and function of islets such as glucose transporter-2,glucokinase,glucagon-like peptide-1 receptor,PDX-1,Ngn3,NeuroDl,Pax-6 and NKX2.2 genes.These islet-like cells might be derived from adult stem cells in bone marrow.

Objective To compare the effects of rat proinsulin gene therapy via intraportal infusion and intramuscular injection blood glucose level in streptozotocin-induced diabetic rots. Methods (1) Recombinant eukaryotic cell expression plasmid of rat proinsulin gene pCMV/proiusulin was transferred into streptozotocin-induced diabetic rats by intraportal infusion and intramuscular injection to observe the effect of rat proiusulin gene therapy in diabetic rats. The treatment group by intraportal infusion (group A) and the group by intramuscular injection (group C) were given pCMV/proinsulin naked plasmid DNA 100 μg, while the control groups by intraportal infusion (group B) or by intramuscular injection (group D) were treated with similar amount of pCMV DNA. Normal group and diabetes mellitus group were also observed at the same time. (2) Blood glucose level was tested and serum insulin was determined by radioimmunoassay. RT-PCR and immunohistochemistry were used to detemine proinsulin mRNA and protein expressions in liver and skeletal muscle and protein. Results (1) The blood glucose levels in two treated groups were both decreased. In group A, levels of blood sugar decreased about 7 mmol/L and glycemie control was maintained for 3-4 weeks. Serum insulin levels step up significantly after pCMV/proinsulin gene therapy. The blood glucose level in group A was significantly lower than those of group B and DM group (P<0.05), while the serum insulin level was higher than those of two groups (P<0.05). In group C, blood glucose levels decreased about 4 mmol/L and glycemic control was maintained for 1-2 weeks. Meanwhile, the concentrations of insulin increased markedly after gene therapy. The blood glucose in group C was significantly lower than those of group D and DM group (P<0.05), while the serum insulin level was higher than those of two groups (P<0.05). (2) Proinsulin mRNA and protein expressions could be detected in either hepatic cell of group A or skeletal muscle cell of group C, not in group B and group D. Conclusion Proiusulin genetherapy via intraportal infusion or intramuscular injection lowers significantly blood glucose in diabetic rats, and thus offers a potential approach to treatment of diabetes.