PLCζ can stably regulate Ca2+ fluctuations in early embryo.

Phospholipase C zeta (PLCζ) is an important inducer of Ca2+ oscillations in mammalian sperm. To explore the influence of PLCζ on early embryonic Ca2+ fluctuations during sperm-egg binding, this study used PLCζ from sheep sperm to construct an early embryonic Ca2+ fluctuation model. First, sheep MII oocytes were cultivated and screened using microinjection technology. Then, a pEGFP-N1-PLCζ plasmid was constructed to activate oocytes in the test group. Ionomycin combined with 6-Dimethylaminopurine (6-DMAP) was used for the control group to explore the effects on early embryonic development and regulation of Ca2+ fluctuations during development. The results demonstrated that both the PLCζ and ionomycin combined with 6-DMAP activation methods induced sheep oocyte parthenogenetic activation and development in early embryos. In comparisons, the cleavage rate of ionomycin combined with 6-DMAP activation was significantly higher than that of PLCζ (60.9% ± 19.4% vs 76.1% ± 0.7%, respectively; p < 0.001), and the blastocyst rates were 16.2% ± 0.62% and 21.1% ± 0.92%, respectively (p < 0.05). Additionally, when comparing the distribution of Ca2+ in early embryos at different stages, Ca2+ in both treatment groups was mainly distributed in the cytoplasm, but the temporal pattern of Ca2+ fluctuations differed. PLCζ resulted in Ca2+ peaks that appeared at the cleavage and morula stages of early embryos, and Ca2+ returned to normal levels at the morula stage. However, the Ca2+ concentration after ionomycin combined with 6-DMAP activation was always much higher than that with PLCζ, and its single peak appeared later than in the PLCζ group. In summary, the PLCζ gene promoted stable regulatory effects on Ca2+ fluctuations at different stages during early embryonic development.

Zinc L-Aspartate enhances intestinal stem cell activity to protect the integrity of the intestinal mucosa against deoxynivalenol through activation of the Wnt/ß-catenin signaling pathway.

The micronutrient, zinc, plays a vital role in modulating cellular signaling recognition and enhancing intestinal barrier function. However, the precise mechanisms underlying the zinc regulation of intestinal stem cell (ISC) renewal and regeneration ability, which drive intestinal epithelial turnover to maintain the intestinal barrier, under physiological and pathological conditions are unknown. In this study, we used in vivo mouse plus ex vivo enteroid model to investigate thoroughly the protection efficacy of zinc L-aspartate (Zn-Asp) on intestinal mucosal integrity exposed to deoxynivalenol (DON). The results showed that 10 rather than 20 mg/kg body weight (BW) Zn-Asp (calculation in zinc) significantly increased the jejunum mass and ameliorated mucosa injury caused by 2 mg/kg BW DON treatment, including improvement of the intestinal morphology and barrier, as well as enteroid-forming and -budding efficiency, which was expanded from crypt cells isolated from jejunum of mice in each group. The repair process stimulated by Zn-Asp was also accompanied by increased fluorescence signal intensity of KRT20 and Villin; increased numbers of MUC2+, CAG+, LYZ+, BrdU+ and Ki67+ cells in mouse jejunum; and protein expression of Ki67 and PCNA in the jejunum, crypt and enteroid. Simultaneously, Zn-Asp increased ISC activity to promote intestinal epithelial renewal even under physiological conditions. These results were further verified in ex vivo enteroid culture experiments, which were treated with 100 µmol/L Zn-Asp (calculation in zinc) and 100 ng/mL DON for 72 h. Furthermore, we demonstrated that Zn-Asp improved intestinal integrity or accelerated wound healing along with Wnt/ß-catenin signaling upregulation or reactivation. Our findings indicate Zn-Asp, especially Zn, enhances ISC activity to maintain the intestinal integrity by activating the Wnt/ß-catenin signaling, which sheds some light upon effective preventive strategies for intestinal injury induced by mycotoxin based on ISCs with exogenous zinc preparations in the proper drugs, health foods or qualified feed.

Methionine and Its Hydroxyl Analogues Improve Stem Cell Activity To Eliminate Deoxynivalenol-Induced Intestinal Injury by Reactivating Wnt/ß-Catenin Signaling.

The intestinal epithelium is derived from intestinal stem cells (ISCs) and has direct contact with nutrients and toxins. However, whether methionine (Met) or a methionine hydroxyl analogue (2-hydroxy-4-(methylthio)butanoic acid (HMB)) can alleviate deoxynivalenol (DON)-induced intestinal injury remains unknown. Mice were treated orally with Met or HMB on days 1-11 and with DON on days 4-8. On day 12, the mice were sacrificed, and the jejunum was collected for crypt isolation and culture. Mouse enteroids were treated with DON and Met or HMB ex vivo. The results showed that Met and HMB increased the average daily feed intake and average daily gain of the mice. Met and HMB also improved the jejunal structure and barrier integrity and promoted ISC expansion, as indicated by the increased enteroid formation efficiency and area, under DON-induced injury conditions. In addition, DON-induced decreases in ISC activity were rescued Wnt/ß-catenin signaling reactivation by Met or HMB in vivo and ex vivo. Collectively, our findings reveal that Met and HMB alleviated DON-induced intestinal injury by improving ISC expansion and reactivating Wnt/ß-catenin signaling. Our study thus provides a nutritional intervention for intestinal diseases involving Wnt/ß-catenin signaling.

Hydrolyzed wheat gluten alleviates deoxynivalenol-induced intestinal injury by promoting intestinal stem cell proliferation and differentiation via upregulation of Wnt/ß-catenin signaling in mice.

Disintegration of the intestine caused by deoxynivalenol (DON), which is a fungal metabolite found in cereal grain-based human and animal diets, triggers severe intestinal inflammatory disease. Hydrolyzed wheat gluten (HWG) can promote the development of intestine. Therefore, HWG was administered orally to male mice on 1-14 days, and DON was administered to them on 4-11 days. Feed, water intake and body weight were recorded all over the experimental period. Blood samples were collected then the mice were sacrificed to collect the jejunum for crypt isolation and culture. The intestinal morphology was observed by electron microscopy, and Western blotting was used to investigate intestinal stem cell (ISC) proliferation and differentiation, as well as the primary regulatory mechanism of the Wnt/ß-catenin signaling. The results showed that HWG increased the average daily gain and average daily water intake of mice under DON-induced injury conditions, and increased the jejunum weight, villous height in the jejunum, and promoted jejunal crypt cell expansion. The DON-induced decrease in Wnt/ß-catenin activity, the expression of Ki67, PCNA and KRT20 were rescued by HWG in the jejunum, crypt and enteroid, as well as the number of goblet cells and Paneth cells. Furthermore, HWG increased jejunum diamine oxidase (DAO) activity. In conclusion, HWG alleviates DON-induced intestinal injury by enhancing ISC proliferation and differentiation in a Wnt/ß-catenin-dependent manner.

Analysis of Characteristics and Relevant Factors of 1 340 Cases of Intentional Injury Cases in Southwest China / 法医学杂志

Objective To investigate the characteristics and patterns of factors such as victims' information, injury tools and time of occurrence of intentional injury cases in southwest China. Methods One thousand three hundred and forty intentional injury cases from several places in southwest China from 2014 to 2016 assessed as minor injury level Ⅱ and above had been randomly selected. Data on victims' information, motives, injury tools, sites of occurrence, time of occurrence, injured parts and degrees of injury were classified and gathered, and then association analyses of motives and types of injury tools as well as degrees of injury and injury tools were made. Results Most of the victims were young adults between 20-50 years （65.2%）, male （82.3%）, rural household registration （62.8%）; the motives were mainly dispute （45.8%）. Injury tools were mostly blunt （54.6%） or sharp （36.0%）. Specifically, injuries were mostly made bare-handed （36.9%） and by cutting tools （33.2%）; the cases mainly occurred in public areas （59.0%）. Cases occurred more frequently in January （11.3%）, February （13.1%）, March （11.6%） and from 22：00 to 01：00 every night. Injuries mainly involved the craniofacial region. The wounds were mainly assessed as minor injury level Ⅱ （61.6%）. There was statistical significance in the difference of types of injury tools among cases with different motives （P<0.05）. There was statistical significance in the difference of the distribution of injury tools among cases with different degrees of injury （P<0.05）. Conclusion The occurrence of intentional injury cases in southwest China has potential patterns and relevant influencing factors. Prevention and analysis of such cases need to be comprehensively considered from the aspects such as victims' information, injury tools and time of occurrence.

Cerebral cavernous malformation 3 gene deficiency promotes early changes in Alzheimer disease-like lesions induced by low lead exposure / 中华预防医学杂志

Objective@#To investigate the effects of cerebral cavernous malformation 3 (CCM3) gene knockout on the lead exposure-induced blood-brain barrier malfunction in mice brain, and the relationship between CCM3 knockout and the Alzheimer's disease (AD).@*Methods@#Wide type (WT) mice and CCM3+/- mice were divided into 4 groups, control group and lead exposed group in WT as well as CCM3+/- mice. Lead exposed groups were treated with 0.05% lead acetate in drinking water for 12 weeks, while control group drink deionized water freely. Blood lead and brain lead levels in each group were detected by graphite furnace atomic absorption spectrometry. The brain tissue of each group was made into paraffin sections, whose morphology were observed by HE staining. The expression of Aβ1-42 in brain tissue was detected by immunohistochemistry and the brain capillaries were labeled by VRGFR2. The protein expression of Claudin-5, ZO-1, and p-Tau was detected by Western blot. The brain tissue RNA was extracted and the relative expression of LRP-1 mRNA was detected by qRT-PCR.@*Results@#The levels of blood lead WT (216.07±84.16) and CCM3+/- (189.64±101.86) μg/L in lead exposed group were higher than those in control group WT (19.52±11.46) and CCM3+/- (11.79±8.20) μg/L, the difference was statistically significant (t=4.18, P=0.006; t=3.79, P=0.016). The levels of brain lead WT (1.78±0.69) and CCM3+/- (1.74±0.66) μg/L were higher than those in control group WT (1.06±0.87) and CCM3+/- (0.97±0.64) μg/L, the difference was statistically significant (t=3.67, P=0.018; t=3.88, P=0.015). The HE staining showed no obvious lesions in the brain of each group of mice. The results of immunohistochemistry showed that there was no Aβ1-42 deposition in the brain of mice in each group. The numbers of microvessels in the brain of CCM3+/- mice in the lead exposed group were decreased. Compared with the relative expression levels of Claudin-5 (WT: 1.30±0.03, CCM3+/-: 1.07±0.08) in control group mice brain, the relative expression of Claudin-5 (WT: 0.96±0.04, CCM3+/-: 0.59±0.01) was decreased with statistical significance (F=199.27, P<0.001). The relative expression level of LRP-1 gene mRNA in brain of lead exposed group (WT: 0.32±0.10, CCM3+/-: 0.06±0.01) was higher than that of unexposed group (WT:1.00±0.06, CCM3+/-:2.12±0.18), the difference was statistically significant (F=288.29, P<0.001). The relative expression level of LRP-1 gene mRNA in brain of CCM3+/- mice exposed to lead was lower than that of WT mice ((0.06±0.01)vs(0.32±0.10), t=26.90, P<0.001).@*Conclusion@#The mice did not show significant AD-like lesions under low-does lead exposure, but resulted in early damage of brain blood-brain barrier and early changes of AD-like lesions in mice, with CCM3+/- mice being sensitive to lead exposure stronger than that of WT mice, suggesting that deletion of CCM3 gene may be one of the potential risk factors for accelerating the development of AD in mice exposed to lead.

The establishment of the immortalized mouse brain microvascular pericytes model and its preliminary application in screening of cerebrovascular toxicants / 中华预防医学杂志

Objective@#To establish the immortalized mouse brain microvascular pericytes model and to apply to the cerebrovascular toxicants screening study.@*Methods@#Brain pericytes were isolated from 3 weeks of mice by tissue digestion. Immortalized pericyte cell line was constructed by infecting with LT retrovirus. Monoclone was selected to purify the immortalized pericyte cell line. The pericyte characteristics and purity were explored by immunocytochemistry. Cell proliferation was measured by using the Pomega MTS cell Proliferation Colorimetric Assay Kit. Pericytes were treated with 0, 160, 320, 640, 1 280, 2 560 μmol/L lead acetate, 0, 5, 10, 20, 40, 80 μmol/L cadmium chloride and 0, 5, 10, 20, 40, 80 μmol/L sodium arsenite in 24 hours. Cell toxicity of each group was determined by MTS assay, median lethal dose (LD50) was calculated in linear regression.@*Results@#Mouse brain pericytes were successfully isolated by tissue separation and enzyme digestion method. After immortalized by LT retroviruses, monoclone was selected and expanded to establish pericyte cell line. The brain pericytes exhibited typical long spindle morphology and positive staining for α-SMA and Vimentin. The proliferation of brain pericytes cell lines was very slowly, and the doubling time was about 48 hours. The proliferation of immortalized brain pericytes cell lines was very quickly, and the doubling time was about 24 hours. After lead acetate, cadmium chloride and sodium arsenite treatment for 24 hours respectively, gradual declines in cell viability were observed. The LD50 of lead acetate was 2 025.0 μmol/L, the LD50 of cadmium chloride was 36.6 μmol/L, and the LD50 of sodium arsenite was 33.2 μmol/L.@*Conclusion@#The immortalized mouse brain microvascular pericyte model is established successfully by infecting with LT retrovirus, and can be applied to screen cerebrovascular toxicants. The toxicity of these toxicants to immortalized mouse brain microvascular pericyte is in sequence: sodium arsenite,cadmium chloride, lead acetate.

Vascular protective effects of aqueous extracts of Tribulus terrestris on hypertensive endothelial injury / 中国天然药物

Angiotensin II (Ang II) is involved in endothelium injury during the development of hypertension. Tribulus terrestris (TT) is used to treat hypertension, arteriosclerosis, and post-stroke syndrome in China. The present study aimed to determine the effects of aqueous TT extracts on endothelial injury in spontaneously hypertensive rats (SHRs) and its protective effects against Ang II-induced injury in human umbilical vein endothelial cells (HUVECs). SHRs were administered intragastrically with TT (17.2 or 8.6 g·kg·d) for 6 weeks, using valsartan (13.5 mg·kg·d) as positive control. Blood pressure, heart rate, endothelial morphology of the thoracic aorta, serum levels of Ang II, endothelin-1 (ET-1), superoxide dismutase (SOD) and malonaldehyde (MDA) were measured. The endothelial injury of HUVECs was induced by 2 × 10 mol·L Ang II. Cell Apoptosisapoptosis, intracellular reactive oxygen species (ROS) was assessed. Endothelial nitric oxide synthase (eNOS), ET-1, SOD, and MDA in the cell culture supernatant and cell migration were assayed. The expression of hypertension-linked genes and proteins were analyzed. TT decreased systolic pressure, diastolic pressure, mean arterial pressure and heart rate, improved endothelial integrity of thoracic aorta, and decreased serum leptin, Ang II, ET-1, NPY, and Hcy, while increased NO in SHRs. TT suppressed Ang II-induced HUVEC proliferation and apoptosis and prolonged the survival, and increased cell migration. TT regulated the ROS, and decreased mRNA expression of Akt1, JAK2, PI3Kα, Erk2, FAK, and NF-κB p65 and protein expression of Erk2, FAK, and NF-κB p65. In conclusion, TT demonstrated anti-hypertensive and endothelial protective effects by regulating Erk2, FAK and NF-κB p65.

Establishment and Evaluation of Hypertensive Rat Model with Excessive Accumulation of Phlegm-dampness Syndrome / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe mainfestations of syndrome and biochemical indices of hypertensive model rats with excessive accumulation of phlegm-dampness syndrome (EAPDS), and to explore its possible pathological mechanism.</p><p><b>METHODS</b>EAPDS rat model was prepared in 50 Wistar rats by feeding with high fat forage. Meanwhile, a normal control group consisting of 10 Wistar rats was set up by feeding with normal forage. After 25-week continuous feeding, 22 rats with body weight (BW) and blood pressure (BP) exceeding 25% those of the control group were selected as a model group. BW, BP, blood lipids, and related serological indicators were detected in all rats. Morphological changes of target organs were observed. mRNA expression levels of leptin receptor (LepR), Janus kinase2 (Jak2), signal transducer and activator of transcription 3 (Stat3), suppressor of cytokine signaling-3 (Socs3), angiotensin II receptor type 1 (AT1), angiotensin II receptor type 2 (AT2), phosphatidylinositol 3 kinase (P13K), serine threonine kinase (Akt), nuclear factor of kappa B (NF-κBp65), inhibitor of nuclear factor kappa-B kinase α (IKKα), NF-kappa-B inhibitor β (lKKβ), NF-kappa-B inhibitor α (IKBα), and AMP-activated protein kinase (AMPK) were detected by quantitative real-time PCR (qPCR). Expression levels of AT1 and LepR in aorta were detected by immunohistochemical assay and Western blot respectively.</p><p><b>RESULTS</b>Compared with the control group, BW, BP, and blood lipids increased; serum levels of leptin (Lep) , Ang II, Hcy, ET-1, TNF-α, IL-6, and p2-MG increased, but NO decreased in the model group (P < 0.05, P < 0.01). Aortal endothelial injury and smooth muscle cell proliferation occurred in the model group, accompanied with heart and renal injury. Compared with the control group, mRNA expression levels of LepR, Jak2, Stat3, Socs3, AT1 , PI3K, Akt, NF-κB p65, IKKβ, IKBα, and AMPK in aorta were up-regulated significantly (P < 0.05), while the expression of IKKa decreased (P < 0.05). Immunohistochem- ical staining showed, brownish yellow deposit of AT1 and LepR was obviously increased, with more extensively positive distribution. Western blot results showed, as compared with the control group, protein expression levels of AT1 and LepR obviously increased in the model group (P < 0.05).</p><p><b>CONCLUSIONS</b>Model rats exhibited typical syndromes of EAPDS. They put up weight with fat abdomen, gloomy hair, poor appetite, hypersomnia, lowered activities , reduced food intake, loose stool, dark red tongue, white tongue with white, thick, greasy fur. Lep could be taken as one of objective indicators for evaluating hypertension rat model with EAPDS.</p>

Aqueous extracts of Tribulus terrestris protects against oxidized low-density lipoprotein-induced endothelial dysfunction / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the role of aqueous extracts of Tribulus terrestris (TT) against oxidized low-density lipoprotein (ox-LDL)-induced human umbilical vein endothelial cells (HUVECs) dysfunction in vitro.</p><p><b>METHODS</b>HUVECs were pre-incubated for 60 min with TT (30 and 3 μg/mL respectively) or 10(-5) mol/L valsartan (as positive controls) and then the injured endothelium model was established by applying 100 μg/mL ox-LDL for 24 h. Cell viability of HUVECs was observed by real-time cell electronic sensing assay and apoptosis rate by Annexin V/PI staining. The cell migration assay was performed with a transwell insert system. Cytoskeleton remodeling was observed by immunofluorescence assay. The content of endothelial nitric oxide synthase (eNOS) was measured by enzyme-linked immunosorbent assay. Intracellular reactive oxygen species (ROS) generation was assessed by immunofluorescence and flow cytometer. Key genes associated with the metabolism of ox-LDL were chosen for quantitative real-time polymerase chain reaction to explore the possible mechanism of TT against oxidized LDL-induced endothelial dysfunction.</p><p><b>RESULTS</b>TT suppressed ox-LDL-induced HUVEC proliferation and apoptosis rates significantly (41.1% and 43.5% after treatment for 3 and 38 h, respectively; P<0.05). It also prolonged the HUVEC survival time and postponed the cell's decaying stage (from the 69th h to over 100 h). According to the immunofluorescence and transwell insert system assay, TT improved the endothelial cytoskeletal network, and vinculin expression and increased cell migration. Additionally, TT regulated of the synthesis of endothelial nitric oxide synthase and generation of intracellular reactive oxygen species (P<0.05). Both 30 and 3 μg/mL TT demonstrated similar efficacy to valsartan. TT normalized the increased mRNA expression of PI3Kα and Socs3. It also decreased mRNA expression of Akt1, AMPKα1, JAK2, LepR and STAT3 induced by ox-LDL. The most notable changes were JAK2, LepR, PI3Kα, Socs3 and STAT3.</p><p><b>CONCLUSIONS</b>TT demonstrated potential lowering lipid benefits, anti-hypertension and endothelial protective effects. It also suggested that the JAK2/STAT3 and/or PI3K/AKT pathway might be a very important pathway which was involved in the pharmacological mechanism of TT as the vascular protective agent.</p>

Expression of leptin receptor and aquaporin 2 in kidneys of obesity-related hyperten-sive rats / 军事医学

Objective To observe the expression of leptin(Lep) receptor (LepR) and aquaporin 2 (AQP2) in the kidneys of obesity-related hypertensive rats ( OHR ) and to explore the mechanism of Lep resistance and water metabolic disorders in them.Methods OHR( model group) were induced by high-fat diet.Normal Wistar rats were chosen as normal control and hypertensive rats(SHR) as positive control.The serum level of triglycerides(TG), total cholesterol(TC), Lep, vasopressin ( AVP ) , angiotensinⅡ( AngⅡ) and β2-microglobulin (β2-MG ) was measured by enzyme linked immunosorbent assays ( ELISA) and renal morphology was observed by HE staining.The density of LepR and AngⅡtype 1( AT1) in the kidney was observed by immunohistochemistry.mRNA And protein expression of LepR and AQP2 in the kidney was assayed by real-time polymerase chain reaction and Western blotting.Results Compared with normal rats,the TG, TC, Lep, AVP, AngⅡand β2-MG of the model group were significantly increased (P<0.05), and protein and mRNA expression of LepR and AQP2 in the kidney were up-regulated (P<0.05).Compared with SHR group, TG, TC and Lep in serum of the model group were significantly increased (P<0.05).The concentrations of AVP,β2-MG and Lep was linearly related(R2 =0.87,R2 =0.95).Conclusion Water metabolic disorder and Lep resistance may be involved in the kidney injury of OHR, which may be one of the important pathogeneses of obesity-related hypertension.

Multipotent differentiation of the EGFP gene transgenic stem cells derived from amniotic fluid of goat at terminal gestational age.

We have isolated stem cells from amniotic fluid of goat at terminal gestational age and transferred the EGFP (enhanced green fluorescent protein) gene into the stem cells previously. The aim of this study was to determine whether the transgenic stem cells have the capability of multipotent differentiation. The transgenic stem cells were induced to differentiate into neurogenic, adipogenic, osteogenic and endothelial cells in vitro. Markers associated with AFS (amniotic fluid-derived stem) cells and the differentiated cells were tested by RT-PCR (reverse transcription-PCR). The results demonstrated that the transgenic AFS cells were capable of self-renewal, a defining property of stem cells. AFS cells were positive for the undifferentiated cell markers, Oct4, Nanog, Sox2 and Hes1, while following differentiation cells expressed markers for neurogenic cells such as astrocyte [GFAP (glial fibrillary acidic protein)] and NSE (neuron-specific enolase), adipogenic cells [LPL+ (lipoprotein lipase+)], osteogenic cells (osteocalcin+ and osteonectin+) and endothelium [CD34+ and eNOS+ (endothelial nitric oxide synthase)]. The results demonstrated that the EGFP gene transgenic AFS cells have the capability of multipotent differentiation, which means that the transgenic AFS cells may be useful in cell-transplantation studies in future.

Differentiation and characteristics of the enhanced green fluorescent protein gene transgenic goat neural stem cells cultured in attached and non-attached plates.

The aims of this study were (i) to determine whether NSCs (neural stem cells) could be isolated from the brain of embryonic day 98 fetal goat, (ii) to determine if these stem cells have the capability of multipotent differentiation following transfection with a reporter gene, EGFP (enhanced green fluorescent protein) and (iii) to study the characteristics of the stem cells cultured in attached and non-attached plates. NSCs were isolated from embryonic day 98 fetal goat brain, transfected with EGFP gene using lipofection, and subcultured in attached and non-attached plates respectively. The transgenic stem cells were induced to differentiate into osteogenic and endothelial cells in vitro respectively. Markers associated with undifferentiated NSCs and their differentiated cells were tested by RT-PCR (reverse transcription-PCR). The results demonstrated that stem cells could be isolated from embryonic day 98 fetal goat brain, and EGFP gene could be transfected into the cells. The transgenic NSCs were capable of self-renewal, a defining property of stem cells, and were grown as free-floating neurospheres in non-attached plates. When the neurospheres were transferred and cultured in attached plates, cells migrate from the neurospheres and are grown as spindle cells. The stem cells were grown as quasi-circular cells when the single stem cells were cultured in attached plates. Both the NSCs cultured in non-attached and attached plates could express Hes1 (hairy and enhancer of split 1), Oct4 (octamer-binding protein 4), Nanog, Sox2 [SRY (sex-determining region Y)-box 2] and Nestin, while following differentiation cells expressed markers for osteogenic cells (Osteocalcin+ and Osteonectin+) and endothelium (CD34+ and eNOS+). The results demonstrated that the goat EGFP gene transgenic NSCs have the capability of multipotent differentiation, which means that the transgenic NSCs may be useful in cell transplantation studies in future.

Differentiation of AFS cells derived from the EGFP gene transgenic porcine fetuses.

We have obtained the EGFP (enhanced green fluorescence protein) gene transgenic porcine fetuses before. The aims of this study were (i) to determine whether stem cells could be isolated from amniotic fluid of the transgenic porcine fetuses, and (ii) to determine if these stem cells could express EGFP and differentiate in vitro. The results demonstrated that stem cells could be isolated from amniotic fluid of the EGFP gene transgenic porcine fetuses and could express EGFP and differentiate in vitro. Undifferentiated AFSs (amniotic fluid-derived stem cells) expressed POU5F1, THY1 and SOX2, while the following differentiation cells expressed markers for chondrogenic (COL2A1), osteogenic (osteocalcin and osteonectin) and neurogenic cells such as astrocyte (GFAP), oligodendrocyte (GALC) and neuron (NF, ENO2 and MAP).

[Effect on the performance of SBBR and the form transformation of nitrogen by different influent pattern].

The difference of sequencing batch biofilm reactor (SBBR) performance and nitrogen transformation mechanism which caused by four different influent patterns were researched. Through variance analysis of SBBR performance, microbial community structure and nitrogen transformation, the results indicated that, on the one hand the dispersed influent pattern displayed higher anti-load ability than the centralized one, under the same efficiency, COD and ammonia load of the dispersed M4 reached 2540 mg x (L x d)(-1) and 540 mg x (L x d)(-1) respectively compared with 2000 mg x (L x d)(-1) and 420 mg x (L x d)(-1) by the centralized M1; on the other hand, considering the dispersed influent pattern, the closer influent mood was to the cycle mood of operation, the higher the nitrogen transformation efficiency was, which finally led residual nitrogen concentration declined.

[Control of shortcut nitrification in SBBR with adequate oxygen supply].

At the high level of dissolved oxygen (DO) in sequencing batch biofilm reactor (SBBR), the approach and mechanism for realizing shortcut nitrification were researched. Landfill leachate was used as handling of object, the mainly environment parameters of the reactor were controlled as follow: DO 5 mg/L, pH 7.0, temperature 25 degrees C, adopted all drainage mode and 12-hour cycle influent. Through mathematical derivation and modeling analysis, determined free ammonia (FA), CO2 and HNO2 as the direct control factors, whereas the influent cycle time was the indirect one, shortcut nitrification was achieved effectively in SBBR. When the volume load of ammonia (NH4(+) -N) was 0.52 kg/(m3 x d) and NaHCO3 was 1.5 mg/L in the reactor, the shortcut nitrification effect was apparent as NH4(+) -N conversion rate was 89% and NO2(-) -N accumulation rate achieved 83% at the same time. With adequate oxygen supply, the key factors of achieving NO2(-) -N accumulation is FA concentration, and as the carbon source of ammonia-oxidizing bacteria, CO2 can upgrade the reactor performance further.

The prevalence of mild cognitive impairment among residents aged 55 or over in Chengdu area / 中华流行病学杂志

<p><b>OBJECTIVE</b>To study the prevalence of mild cognitive impairment (MCI) in the urban and the rural areas in Chengdu, Southwest China.</p><p><b>METHODS</b>Residents aged 55 or over were selected by stratified random cluster sampling from 19 districts, cities, and counties of Chengdu area in Sichuan province. A two-stage survey was carried out. In the first stage, CMMSE, CES-D were used as screening instruments. In the second stage, Diagnostic questionnaires of dementia and CDR were used as diagnostic instruments. The diagnostic criteria of mild cognitive impairment adopted from Petersen's were: (1) memory complaint; (2) normal activities of daily living; (3) normal general cognitive function; (4) memory impairment incompatible with age; (5) not demented; (6) CDR = 0.5 and (7) exclusion of the reversible cognitive impairment caused by other factors (i.e. depression).</p><p><b>RESULTS</b>Three thousand, nine hundred and ten subjects were examined. The prevalence rates of MCI was 2.4%. The MCI prevalence rates in the urban and the rural areas were 1.5%, 2.5% respectively, without significant difference. The MCI prevalence in males and females were 1.8%, 2.9% respectively. Prevalence rate in female was higher than in males with significant difference. Prevalence of illiteracy (4.0%) was the highest among different educational levels. The accumulated prevalence increased with age.</p><p><b>CONCLUSION</b>The prevalence of MCI (2.4%) was slightly higher than the prevalence of AD (2.05%) in the same areas of Chengdu. MCI seemed to be a high risk factor for AD which should to be followed up. Early intervention in MCI might be helpful in the prevention of AD.</p>