Objective We increase the soluble expression of artificial tandem hybrid ubiquitin binding domains ( ThUBD) in prokaryotic cytoplasm of Escherichia coli BL21 ( DE3 ) , which offer an effective and special profiling for ubiquitin conjugates( UbC) .Methods Codon optimization of the ThUBD was performed, followed by analysis of codon relative adaptiveness based on relative frequency of synonymous codon ( RFSC) of E.coli.Further induced expression and yeast ubiquitin conjugate enrichment quantified the soluble ThUBD-S and tested the ability to bind UbC.Results The statistical result showed that the percentage of codon of the highest usage frequency was increased from 48%to 75%, and codon adaptation index( CAI) was increased from 0.63 to 0.88 after codon optimization, which might suggest a higher expression of the ThUBD in E.coli BL21 (DE3).The subsequent SDS-PAGE indicated that the soluble target protein was increased four times, which accounted for 13.06%of total cell lysis.Further ubiquitinated proteome of yeast demonstrated that the ability to bind and enrich UbC of optimized ThUBD-S did not change compared with original ThUBD.Conclusion The expression of ThUBD-S can quadruple after codon optimization.At the same time, codon optimization does not impact its soluble expression and the ability to bind UbC.

BACKGROUND: In the central nervous system(CNS) of normal adults there are neural stem cells or neural progenitor cells, which are capable of self-renewing and multiple differentiating. In normal physiological conditions, intrinsic neural stem cells are in a resting state. What state will they be in during cerebral ischemia?OBJECTIVE: To observe the distribution, proliferation and differentiation of intrinsic neural stem cells in focal transient ischemia in rats.DESIGN: A randomized controlled exploratory trial based on the rats.SETTING: Neurobiological department, histological and embryological department of a medical college.MATERIALS: The experiment was conducted in the Neurobiological Department of Luzhou Medical College from July 2001 to July 2002. Altogether 41 healthy adult SD rats of either gender, weighting 250 g - 300 g, were selected.INTERVENTIONS: The focal ischemia model was made by blocking middle cerebral artery(MCA) and reperfusing for 0.5 hour, 3 hours, 6 hours, 12 hours, 1 day, 2 days, 3 days, 5 days and 10 days. Sham-operation group was treated by the same method, but the filament was not long enough to block MCA, and normal rats served as control group. The rats were sacrificed at given time points, and their brains were made into cerebral slices. The single-and double-labeled immunohistochemical staining was employed to detect the proliferation, distribution and differentiation of intrinsic neural stem cells.MAIN OUTCOME MEASURES: The distribution, proliferation and differentiation of intrinsic neural stem cells.RESULTS: Immunoreactivity of proliferating cell nuclear antigen(PCNA)was present in most ependymal cells in ventricular zone(VZ), and PCNA-positive cells were sparsely distributed in the parenchyma in normal and sham-operation groups. At 3 hours of reperfusion, PCNA-labeled cells were first detected in rostral subventricular zone. At 12 hours of reperfusion and onward, PCNA-positive cells appeared in some choroid plexus cells in bilateral lateral VZ. At day 3 to day 10 of reperfusion, PCNA-labeled cells significantly increased in infarct boundary in preoptic area, striatum and deep layer of frontoparietal cortex. PCNA-labeled cells were first detected in subgranular zone of dentate gyrus 3 days after reperfusion, and increased with time. A very small number of double-positive cells expressed with PCNA and glial fibrillary acidic protein(GFAP) were first detected in infract boundary in preoptic area on day 3 and onward. No double-PCNA and NF-positive cells were detected within 10 days of reperfusion.CONCLUSION: Focal cerebral ischemia activates intrinsic neural stem cells, which proliferate and differentiate, and migrate toward ischemic striatum and frontoparietal cortex. This may help clarify the mechanism of functional recovery after ischemia.

AIM:To evaluate the efficacy and safety of large sized continuous curvilinear capsulorhexis ( CCC ) involving zonular area in manual small incision cataract surgery ( MSICS) .
METHODS:Totally 1 443 cataract patients ( 1 965 eyes ) underwent MSICS, in which large CCC up to 7-8mm were performed . The related conditions such as success rate of performing CCC, capsule rupture, corneal edema and visual acuity after surgery were recorded and statistically analyzed.
RESULTS:In total of 1 965 eyes, 1 942 eyes ( 98. 83%) had successful CCC, 15 eyes (0. 76%) had a radial tear which could not be saved when performing CCC, 8 eyes (0. 41%) had anterior capsule opened with a cystotome or a scissor due to capsule membrane calcification. No posterior capsule rupture occurred, and intraocular lens was transplanted in all cases. Transient corneal edema was noted in 36 eyes (1. 83%). One day after surgery, 1650 eyes (83. 97%) had visual acuity ≥0. 5, 1867 eyes (95. 01%) ≥0. 3. No unstable intraocular lens was noted in all cases.
CONCLUSION:Large CCC technique for MSICS is safe and reliable, not decreasing stability capsular bag and intraocular lens. It is of high originality despite zonular area is involved so that it is worthy of application in basic medical institutions.

Objective To investigate the com and pepper fluorine in Zhenxiong county of Yunnan province, as well as the change of com and pepper fluorine after baked by coal, clay-mixed with cual for a relatively long-term, in order to provide a scientific basis for reducing fluorine intake. Methods The endemic areas of Yile, Wufeng's Songlinwan, Tangfang and Wufeng's Wugu in Zhenxiong county, and a non-endemic area Xiaguan in Dali city were selected as study sites. Ten samples of fresh corn and pepper were collected in each region, and fluorine was determined using acid leaching/potentiometry freshly and after baking or drying for 10 days or 4 months, respectively. Results The fluorine content of local fresh corn in Xiaguan of Dali city and Yile,Wufeng's Songlinwan, Tangfang, Wufeng's Wugu in Zhenxiong county were (1.31 ± 0.13),(1.65 ± 0.64),(1.92 ±0.37), (2.32 ± 0.49), (1.98 ± 0.66)mg/kg, respectively, and there were statistically significant differences across the regions(H = 27.871, P ＜ 0.05). The fluorine content of corn samples after baking or drying for 4 months were ( 1.82 ± 0.17), (26.43 ± 12.03), (39.27 ± 8.09), ( 14.27 ± 4.37), ( 14.33 ± 1.73)mg/kg, respectively, which were significantly higher than that of the fresh com in the corresponding region(all P ＜ 0.05 ), and there were statistically significant differences across the regions(H = 42.512, P＜ 0.05). The fluorine content of the local fresh chili were (3.34 ± 1.08), (3.44 ± 0.55), (3.47 ± 0.74), (3.46 ± 0.93)mg/kg, respectively, in the 4 observed places in Xiaguan of Dali city and Yile, Wufeng's Songlinwan, Tangfang in Zhenxiong county, and there were no statistically significant differences across the regions (F = 0.052, P ＞ 0.05 ). The fluorine content of pepper samples after baking or drying for 4 months were (7.01 ± 1.64), (226.07 ± 83.69), (179.36 ± 148.37), (54.51 ± 34.67)mg/kg,respectively, which were significantly higher than that of the fresh pepper in the corresponding region(all P ＜ 0.05 ),and there were statistically significant differences across the regions(H = 28.822, P ＜ 0.05). Conclusion Corn and chili fluorine is significantly increased after baked with coal and clay-mixed with coal by farmers in Zhenxiong county, a coal- burning borne endemic fluorosis area of Yunnan province.

Objective:To prepare a novel dental lithium disilicate glass-ceramic,and to study its properties.Methods:A2 colored novel glass-ceramic was prepared by sintering technique.The flexural strength,density,elastic modulus,hardness,fracture toughness, translucency and microstructure of the novel glass-ceramic and IPS e.max Press (LT A2,MO1 and HO1 )ceramic ingots were studied. Results:The novel glass-ceramic demonstrated a lower flexural strength (31 5 MPa)than MO ceramic ingot(338 MPa)(P <0.05). The other mechanical properties of the novel glass-ceramic were similar to those of LT and MO ceramic ingots(P >0.05).The translu-cency parameter of the novel glass-ceramic(21 .2)was higher than that of HO(1 6.5)but lower than that of LT(27.8)and MO(27.5) ceramic ingots(P <0.05).SEMimages showed an interlocking microstructure of rod-shaped Li2 Si2 O5 crystals in all the glass-ceramics investigated.Conclusion:The mechanical properties,translucency and microstructure of the novel glass-ceramic are similar to those of IPS e.max Press ceramic ingots,which can meet the requirements of clinical application.

Objective To investigate the relationship and mechanism of the serum vitamin D levels with insulin sensitivity and adipokines APN,RBP4,visfatin and TNF-α in patients with gestational diabetes mellitus(GDM). Methods A total of 101 patients with gestational diabetes( GDM group) and 50 pregnant women with normal blood sugar( control group) were recruited to detect the serum levels of correlative biochemical indexes,APN,RBP4,visfatin,TNF-α and 25OHD3 using an HPLC method. Twenty-four patients in the GDM group diagnosed with vitamin D deficiency(<25 nmol·L-1 ) were randomized to receive either 1,25(OH) 2D3(0. 25 μg·d-1) orally or no treatment. Serum levels of correlative biochemical indexes were detected after 1 month. Results GDM patients had 25OHD3 levels significantly lower than the control group(P<0. 05),APN levels lower than the control group,and RBP4,visfatin,TNF-α levels significantly higher than the control group(all P<0. 05). After 1 month of 1,25(OH) 2D3 treatment,HOMA-IR increased,while HOMA-β decreased. APN levels was positively correlated(r= 0. 526) with 25OHD3;RBP4,visfatin,TNF-α decreased and were negatively correlated with 25OHD3( r values were -0. 272,-0. 153,-0. 072). Conclusion Vitamin D can reduce the adipokines RBP4,visfatin,TNF-α,increase APN,and thus play a protective role for gestational diabetes.

Previous experiments has shown that Ginsenoside Rb1 (GRb1), which is one of the most important active ingredients in ginseng (Panax ginseng C.A. Meyer), reduced infarct and neurologic deficit followed by the transient cerebral ischemia in rats. The mechanism of this neuroprotective function is unclear. In this study, we tested whether the neuroprotective effect of GRb1 is achieved through preventing the neuronal apoptosis and modulating expression of neuronal apoptosis inhibitory protein (NAIP). Focal cerebral ischemia was induced by the middle cerebral artery occlusion (MCAO) in Wistar rats. GRb1 (40 mg/kg, i.p.) was administered immediately after the onset of reperfusion. The rats with neurological deficits were randomly divided into 2 groups: the ischemia and the GRb1 group. Each group was again divided into subgroups according to the various reperfusion time (3 h, 12 h, 1, 2, 3, 5, 10 days, n=4 per time point). Apoptotic cells were analyzed using TUNEL. Immunohistochemical method was used to assess expression of NAIP. This results showed that the number of apoptotic cells elevated at 3 h of reperfusion, and peaked at 24 h, then declined, but the number of apoptotic cells at 10 d after ischemia was significantly more than those of control groups (P＜0.01). Compared with ischemia group, the apoptotic cells decreased at all subgroups of GRb1; however, the significant differences were only found from 12 h to 3 d of reperfusion. In normal and sham groups, NAIP weak immunostaining was diffusely present in the neurons of parenchyma. The number of NAIP-positive cells started to increase in ischemic regions at 3 h after ischemia, peaked at 12 h and declined up to 5 d of reperfusion. At 5 d after ischemia, the number of NAIP-positive cells was less than that of control group (P＜0.05). A few astrocytes strongly expressed NAIP in the ischemic area. In the GRb1 group, the number of NAIP-positive cells from 12 h to 10 d after ischemia was evidently higher than in the ischemia group. Thus, these results suggest that GRb1 has potential ability to prevent apoptosis, the mechanism of which is related to induce expression of NAIP.

BACKGROUND:Previous studies have demonstrated that cell transplantation has neuroprotective effect on intracerebral hemorrhage,and some researches have indicated that transplantation of bone marrow stromal cells(BMSCs)can promote neural function recovery after cerebral infarction.OBJECTIVE:To explore whether transplantation of BMSCs-modified by gtial cell line-dedved neurotrophic factor gene(GDNF)gene provides a better therapeutic effect than native BMSCs after stroke.METHODS:Totally 36 SD rats were induced intracerabral hemorrhage models by injecting autologous arterial blood,and then divided into 3 groups(n=6).each group was assigned into 2 sub-groups Rabbits in each group were stereotaxically grafted with 20 μL GDNF/BMSCs,BMSCs or saline respectively.The rats were executed at 1 and 2 weeks after operation,and immunohistochemistry was used to observe the expressions of synaptophysin(Syn)and growth associated protein-43(GAP-43)in the margin of the hemorrhagic focus.RESULTS AND CONCLUSlON:Compared with the BMSCs and control groups.both Syn-immunoreactive and GAP-43-immunoreactive products were significantly increased in the GDNF/BMSCs group(P＜0.05).Present results demonstrate that transplantation of GDNF gene-modified BMSCs provides better neuroprotection than native BMSCs delivery for stroke.

Objective To observe the incidence of fragmented QRS complex (fQRS)and ST Segment depression fQRS (STD fQRS)during the first 48 hours after non-ST elevation myocardial infarction(NSTE MI)and discuss the value of predicting mortality in patients with NSTE MI .Methods Based on the ECGs ,the patients with NSTE MI were divided into two groups :fQRS and non fQRS group .And then fQRS group was divided into two sub-groups :STD fQRS and non-STD fQRS group .Their mortality was studied during long-term follow-up .Results (1)731 patients with NSTE ACS [the NSTE MI group(n=609) and the UA group(n=122)] were studied .The incidence of fQRS in the NSTE MI group was higher than that of the UA group .(2)All cause mortality in the fQRS group were higher than that in the non-fQRS group ,and all-cause mortality in the STD fQRS group were higher than that in the non-STD fQRS group ,all the above results were not only in the early stages of NSTE MI ,but also in the long term fol-low-up .(3) Multivariate Cox regression analysis revealed that STD fQRS was an independent significant predictor for all cause mortality ,but not of the fQRS .Conclusion The STD fQRS may be an independent predictor of mortality in patients with NSTE MI .

Objective To investigate the pathogenesis of peripheral blood mononuclear cells(PBMC) nuclear factor kappa-?B(NF-?B) in children with primary nephritic syndrome(PNS) and the effect of astragalus on the activity of NF-?B.Methods Twenty-five children with PNS and 20 normal children were studied.Isolated PBMC were separated from 5 mL venous blood in asepsis condition.NF-?B stimulator,NF-?B inhabitor and astragalus were added into the different tubes of PBMC,respectively.The nuclear protein was extracted from the pellets and the optical density(A) values of nuclear protein was measured by enzyme-linked immunosorbent assay(ELISA).Results The activity of PBMC NF-?B in PNS group was higher than that in normal group(P0.05).Astragalus could decrease the activity of PBMC NF-?B which had been stimulated by interleukin-1?(IL-1?)(P