Cyclic peptide drugs have gradually become an emerging research direction due to their some favorable properties such as high-efficiency binding affinity, high selectivity, lower toxicity, and stable metabolism. In recent years, the number of cyclic peptide drugs under clinical research has continued to increase. Unlike the previous cyclic peptide drugs, which were mostly derived from natural products and their derivatives, these cyclic peptide drugs are designed by genetically encoded display technologies which are based on rational design and in vitro evolution (such as BT1718, PTG-300, POL6326, etc). Among them, phage display technology has some advantages such as mature research system, low cost, and simpler operation that make it well recognized and praised by the majority of researchers in this field. Here, we reviewed the recent progress of applying phage display technology to explore diverse cyclic peptide libraries, which, we believe, will contribute more valuable candidate cyclic peptide drugs in clinical research.

OBJECTIVETo observe the intervention effects of electroacupuncture at "Yishu" (EX-B 3) on rats with type-2 diabetes mellitus (T2DM), so as to provide experiment references for acupuncture to treat T2DM.

METHODSAmong seventy male Wistar clean-grade rats, 8 rats were randomly selected into a control group; the rest rats were made T2DM model. Fifty-two rats which were successfully made T2DM model, according to randomized block method, were divided into a model group (10 rats), a medication group (10 rats), an electroacupuncture at "Shenshu" (BL 23) group (11 rats), an electroacupuncture at "Pishu" (BL 20) group (10 rats) and an electroacupuncture at "Yishu" (EX-B 3) group (11 rats). Seven days after successful establishment of model, the rats in the model group were fixed in the self-made rat bag without receiving any treatment; the rats in the medication group, according to body mass (10 mL/kg), were treated with intragastric administration of glimepiride; the rats in all the electroacupuncture groups were treated with electroacupuncture at "Shenshu" (BL 23), "Pishu" (BL 20) and "Yishu" (EX-B 3), respectively. The continuous wave was selected with a frequency of 15 Hz and a current intensity of 4 to 6 mA. The treatment was given 20 min per treatment, once a day, 5 treatments per week for continuous 4 weeks. Before the establishment of model and continuous 4 weeks after the intervention, blood samples were collected from rats' caudal vein, and fasting blood glucose (FBG) was measured with FBG device each week. After the last intervention, the rats were killed and hypothalamus, pituitary and adrenal gland were collected. The colorimetric method was applied to measure the contents of triglyceride (TG), cholesterol (TC), high-density lipoprotein (HDL-C) and low-density lipoprotein (LDL-C); radioimmunoassay was used to test the contents of glycated serum protein (GSP), fasting insulin (FINS), corticotropin-releasing hormone (CRH), adrenocorticotropic hormone (ACTH) and cortin (CORT).

RESULTSFour weeks after the intervention, except that the rat's body mass in the normal group continued to increase, body mass in the model group, medication group and each electroacupuncture group were significantly reduced (P<0.01, P<0.05). Compared with the model group, the FBG in the electroacupuncture at "Pishu" (BL 20) group and electroacupuncture at "Yishu" (EX-B 3) group were obviously reduced (P<0.05, P<0.01); FBG in the electroacupuncture at "Yishu" (EX-B 3) group was lower than that in the medication group and electroacupuncture at "Shenshu" (BL 23) group (both P<0.05). The contents of TG, HDL-C and LDL-C in the electroacupuncture at "Yishu" (EX-B 3) group were reduced (P<0.05, P<0.01), the content of TG was significantly lower than that in the medication group and electroacupuncture at "Shenshu" (BL 23) group (both P<0.05), the content of LDL-C was significantly lower than that in electroacupuncture at "Shenshu" (BL 23) group (P<0.05). Insulin sensitivity index (ISI) in the medication group, electroacupuncture at "Pishu" (BL 20) group and electroacupuncture at "Yishu (EX-B 3)" group were evidently increased (P<0.05, P<0.01); ISI in the medication group was lower than that in the electroacupuncture at "Yishu" (EX-B 3) group (P<0.05). The content of CRH in the electroacupuncture at "Yishu" (EX-B 3) group was lower than that in the medication group and electroacupuncture at "Shenshu" (BL 23) group (P<0.05, P<0.01); the content of CORT in the electroacupuncture at "Yishu" (EX-B 3) group was lower than that in the medication group and electroacupuncture at "Pishu" (BL 20) group (both P<0.05).

CONCLUSIONElectroacupuncture at "Yishu" (EX-B 3) could reduce the level of CORT to improve the insulin resistance in rats with T2DM, improve insulin sensitivity index, regulate blood lipid metabolism and relieve the hyperactivity of the HPA axis.

Acupuncture Points ; Acupuncture Therapy ; Animals ; Blood Glucose ; metabolism ; Diabetes Mellitus, Type 2 ; metabolism ; therapy ; Electroacupuncture ; Hormones ; metabolism ; Humans ; Hypothalamus ; metabolism ; Male ; Pituitary-Adrenal System ; metabolism ; Rats ; Rats, Wistar

Objective To investigate the effects of berberine on tumor-associated macrophages (TAM)and the expression of cyclooxygenase-2 (COX-2)of intestinal polyps in Apc(Min/+) mice.Methods A total of 20 Apc(Min/+) mice,four weeks old,were equally divided into the control group and the berberine group,10 in each group.The mice of the control group drank plain water,while the mice of berberine group drank water with 0.1 % berberine.After 12 weeks,all the mice were sacrificed.The intestine and colon were isolated,and the numbers of polyps were counted.The expression of F4/80,inducible nitric oxide synthase-2 (iNOS),macrophage mannose receptor (MR)and COX-2 was detected by immunohisto-chemistry method.The relative expression of COX-2 at protein level was measured by Western blotting. The t test was performed for comparison between two independent groups.Results The total number of intestinal polyps,the number of small intestinal polyps and the number of colon polyps of the berberine group (11 .50±2.05 ,10.50±1 .77 and 1 .00±0.46,respectively)were all less than those of the control group (30.63±1 .69,28.00±2.00 and 2.63±0.74,respectively),and the differences were statistically significant (t=16.727,16.952 and 3.162,P =0.001 ,0.001 and 0.010,respectively).The percentage of F4/80 positive cells in the stroma of polyps of the berberine group ((17.40 ±4.23 )%)was less than that of the control group ((31 .24±6.34)%),and the difference was statistically significant (t =5 .327, P =0.043).The percentage of iNOS positive cells in the stroma of polyps of the berberine group ((7.43± 1 .78 )%) was higher than that of the control group ((2.72±0.68)%), and the difference was statistically significant (t=7.335 ,P =0.004).The percentage of MR positive cells in stroma of polyps of the berberine group ((19.52±1 .54)%)was less than that of the control group ((12.63±0.68)%),and the difference was statistically significant (t=5 .634,P =0.016).The percentage of COX-2 positive cells in stroma of polyps of berberine group ((3.38 ± 0.51 )%)was less than that of the control group ((7.60±0.57 )%),and the difference was statistically significant (t = 7.234,P = 0.001 ).The relative expression of COX-2 at protein level of polyps of the berberine group was lower than that of the control group. Conclusion Berberine may take the role in inhibiting the growth of intestinal polyps in Apc(Min/+) mice through interfering the differentiation of TAM in polyps and suppression the expression of COX-2.

Pseudorabies virus (PRV) is a swine herpesvirus of the Alphaherpesvirinae subfamily and a pathogen of swine resulting in devastating disease and economic losses worldwide. Cre/loxP site-specific system has the character of site specific, time specific, tissue specific and high efficiency in recombination, which makes this system universal in vivo and in vitro recombination of bacteria, fungus, plants, insects and mammals. A recombinant PRV which contain a loxP site in TK locus by using Cre/LoxP recombinant system was construsted. A pair of primers were synthesized according to the pEGFP-C1 sequence published on GenBank, and were used to amplify the EGFP gene expression cassette with two loxP sites flanking each side. This target gene was cloned into pSKLR, the resulting transfer vector pSKLR-GFP-loxP was then cotransfected into 293T cells with PRV SH strain genomic DNA. The recombinant virus rPRV1 was selected and purified in TK-143 cells by choosing fluorescent expressing plaques. Cre expression vector pOG231 was cotransfected into 293T cells with rPRV1 genomic DNA. The second recombinant virus rPRV2 was obtained, which contains only one loxP site in TK locus. Sequencing results of rPRV2 TK gene indicated that 34bp loxP site was inserted into rPRV2 genome and there were 270bp deletion in TK gene. PCR amplifying different generations of rPRV2 TK gene showed that the mutant was stable when passages in RK-13 cells. TCID_ 50 assay indicated that rPRV2 grows well on RK-13 cells. The LD_ 50 test results on BALB/C mice suggested that the virulence of rPRV2 was reduced. As a conclusion, the report gene GFP expression cassette was removed successfully from rPRV1 genome and only one LoxP site was leaved in rPRV2 genome by using Cre/LoxP recombinant system.

Amblyopia greatly affects the physical and mental development of children. Acupuncture is effective for amblyopia, though its mechanism remains unclear. This article summarized the mechanism of acupuncture treatment of amblyopia from the perspectives of morphology of neurons in visual cortex, visual electrophysiology, and molecular biology, etc. It was found that acupuncture may treat amblyopia through repairing the morphological and ultrastructural damages of neurons in visual cortex, promoting the electrical activities in visual pathway and visual cortical neurons, and modulating the synthesis and expression levels of factors involved in visual system. Nevertheless, further studies are required to unveil the mechanism of acupuncture treatment of amblyopia.

We designed a locator for puncture under CT examination.Thirty-three patients of both sexesaged 16-76 yr weighing 46-80 kg undergoing bilateral thoracic or lumber sympathetic nerve block under CT examination were enrolled in this study.One side was punctured under the guidance of the locator,while the other side was punctured by conventional technique.The time consumed during puncture,CT scan adjusting time,the deviation of the needle from the targets and the incidence of puncture complications were recorded.The results showed that compared with the conventional technique,with the locator the time consumed during puncture was significantly shorter,the number of attempts and the deviation of the needle from the target were significantly reduced.The procedure was successfully performed on both side in all patients without serious complications such as hemothorax and pneumothorax.

Objective To investigate the value of three-dimensional speckle tracking imaging (3D-STI) in assessment of left ventricular (LV) strains. Methods Thirty healthy young adults examined by two-dimensional speckle tracking imaging (2D-STI) and 3D-STI. And the results of LV measurements were compared, which included mean peak systolic longitudinal strains, radial strains and circumferential strains. Also, the time consumption of these two methods was compared. Results The time needed for 3D-STI in acquisition and analysis of the images were (309.3±23.4)s, (305.5±11.2)s, while the time for 2D-STI were (490.6±14.4)s, (1261.4±39.9)s. The differences were signiifcant(t=-21.81, 69.94, both P<0.01). The global mean peak systolic radial strains was (48.59±7.68)%by 3D-STI and (33.25±7.27)%by 2D-STI. The difference was signiifcant(t=9.16, P<0.01). The global mean peak systolic longitudinal and circumferential strains were (-17.66±3.14)%, (-17.13±2.29)% by 3D-STI and (-21.35±2.46)%, (-21.97±3.84)% by 2D-STI. The differences were signiifcant(t=5.33, 5.99, both P < 0.01). The 3D-STI strains were different at different levels of LV. The longitudinal, circumferential and radial 3D-STI strains were largest at middle levels. However, 2D-STI strains didn′ t show such trend. Peak strains measured by 3D-STI and 2D-STI showed high inter-observer and intra-observer agreement in Bland-Altman chart. Conclusion 3D-STI is a novel, convenient and reproducible method to evaluate the strains of LV.

OBJECTIVETo evaluate the feasibility of using polyporus silk fibroin as a kind of novel material for facial nerve regeneration.

METHODSThe porous silk fibroin conduit was used in the reconstruction of a 5 mm facial nerve gap of SD rat. Chitosan conduit was taken as control group. General observation, electrophysiological study, histological study and image analysis were performed 2, 4, 6 and 8 weeks postoperatively.

RESULTSThe facial nerve of SD rat regenerated successfully as time passed through. Mean CAP percentage of regenerated nerve in SF conduit was 24.94% +/- 5.73% 8 weeks postoperatively, which had no statistical significance with that of chitosan conduit group (P = 1.125). And the average number of myelinated myelinated nerve fibers in SF conduit was 62. 5 +/- 6. 3, which had statistical significance with that in chitosan conduit group (P = 0.016).

CONCLUSIONSThe porous silk fibroin conduit could effectively repair facial nerve defect and improve peripheral nerve functional recovery.

Animals ; Facial Nerve Injuries ; surgery ; Female ; Fibroins ; pharmacology ; therapeutic use ; Materials Testing ; Nerve Regeneration ; drug effects ; Rats ; Rats, Sprague-Dawley ; Wound Healing

OBJECTIVETo explore the effect of clinical application of stand-alone MC+PEEK cage in anterior cervical fusion.

METHODSFrom January 2011 to January 2014,50 patients were treated with the MC+PEEK cage filled with autogenous cancellous illic-bone graft after anterior cervical discectomy. There were 22 patients with cervical spondylosis,26 patients with traumatic cervical disc herniation, 2 patients with cervical instability in these patients. There were 32 males and 18 females, aged from 30 to 79 years old with an average of 53.30 years old. There were 32 patients with single segment, 15 patients with double segments and 3 patients with three segments. Cervical AP and lateral and the flexion-extension X-rays were regularly taken in order to assess the cervical physiological curvature, the graft fusion and internal fixation related complications. Nerve function, clinical effect and bone fusion were respectively evaluated according to Japan Orthopedic Association (JOA), Otani grade and Suk method.

RESULTSAll patients were followed up from 6 to 36 months with an average of 20 months. No correlated surgical complications were found and all patients obtained bony fusion with an average time of 4.30 months. JOA score had significantly improvement after surgery (P < 0.05). The JOA score was 10.60 ± 3.00 before surgery and 16.10 ± 2.20, 16.40 ± 2.35 at one week and six months after surgery respectively. According to Otani grade,40 cases got excellent results, 9 good, 1 fair. No significant dysphagia and internal fixation related complications such as displacement of cages were found during the follow-up period.

CONCLUSIONUsing this cage in anterior cervical fusion can obtain satisfactory clinical effect with less operation injury and reduce the complications. It is a better fusion method in anterior cervical fusion.

Adult ; Aged ; Cervical Vertebrae ; surgery ; Female ; Humans ; Male ; Middle Aged ; Spinal Fusion ; adverse effects ; instrumentation ; methods

Objective Transcription factor forkhead box L 2 (FOXL2) is a key regulator of granulosa cells (GCs) estrogen syn-thesis and function maintenance .However, the FOXL2 protein expres-sion and function regulation mechanism are unknown .We explored how DNAJB11 regulates estrogen synthesis of granulosa cells with immunoprecipitation , immunofluorescent staining and luciferase re-porter gene. Methods The expression and localization of DNAJB 11 was detected by immunohistochemistry staining in isolated mouse ovary tissues .we use immunoprecipitation , immunofluorescence staining and luciferase reporter gene assay to investigate the mechanism of DNAJB11, a member of the endoplasmic reticulum Hsp 40 /DnaJ family, regulating the estrogen synthesis in granulosa cells . Results DNAJB11 is expressed in the mouse ovary and granulosa cells .Follicle-stimulating hormone (FSH) promotes DNAJB11 ex-pression in a time and concentration dependent manner and induces endogenous DNAJB 11 protein translocation from the ER to the nu-cleus in KGN cells.Moreover, Adenovirus-mediated overexpression of DNAJB11 did not affect the proliferation of granulosa cells .How-ever, the concentration of estrogen in granulosa cells was affected by concentration -dependent and subcellular localization-dependent manner (10749±801.7 pg/mL vs 14217±1218.0 pg/mL P<0.01).Immunoprecipitation assay confirmed that DNAJB11 binds to FOXL2 in granulosa cells .When overexpressed in the nucleus of granulosa cells , DNAJB11 could significantly enhance the stability of FOXL2 (P<0.05) and promote FOXL2-mediated activity of Cyp19A1 promoter (P<0.01), while the expression of DNAJB11 in the nucleus increased the expression of Cyp 19A1 protein by 1.5 times ( P<0.05) . Conclusion These results demonstrate that DNAJB 11 was a new binding molecule of transcription factor FOXL 2 and regulated FOXL 2 protein stability and transcription activity .