[The effect and mechanism of exosomes derived from human amniotic epithelial cells on the proliferation and migration of HaCaT in high glucose environment].

Objective: To investigate the effect and mechanism of exosomes derived from human amniotic epithelial cells (hAEC-Exos) on the proliferation and migration of HaCaT in high glucose environment. Methods: The experimental research method was adopted. The amniotic membrane tissue was collected from 10 healthy pregnant women at full term delivery in the Department of Obstetrics and Gynecology of Fujian Medical University Union Hospital from January to June 2019, and the primary human amniotic epithelial cells (hAECs) were isolated. The growth status and morphological changes of the primary hAECs on the 2nd, 4th, and 7th day of culture were observed, and the expressions of the cells surface markers of CD73, CD90, CD29, CD34, and human leukocyte antigen DR (HLA-DR). The 2nd to 4th passages of hAECs were used in the following experiments. The hAEC-Exos were separated by ultracentrifugation method. The HaCaT and hAEC-Exos were co-cultured for 3 h, and the uptake of hAEC-Exos by HaCaT was observed by inverted fluorescence microscopy. The HaCaT were divided into phosphate buffer solution (PBS) group and hAEC-Exos group or dimethyl sulfoxide (DMSO)+PBS group, DMSO+hAEC-Exos group, and LY294002+hAEC-Exos group, which were dealt correspondingly, with 3 wells in each group. Cell counting kit 8 (CCK-8) method was used to detect cell proliferation activity after 0 (immediately), 12, 24, 36, 48, and 60 h of culture. The scratch test was conducted to detect the scratch healing at 0, 24, 48, and 72 h after the scratch, and the scratch healing rate was calculated, respectively. The Transwell experiment was conducted to detect the number of transmembrane cells after 48 h of culture. The Western blotting was used to detect the protein expressions of mammalian target of rapamycin (mTOR), phosphorylated mTOR (p-mTOR), protein kinase B (Akt), and phosphorylated Akt (p-Akt) related to phosphatidylinositol 3-kinase-Akt-mTOR (PI3K-Akt-mTOR) pathway after 24 h of culture. Data were statistically analyzed with analysis of variance for repeated measurement, one-way analysis of variance, and independent sample t test. Results: Most of the primary hAECs were oval and uniform in size on the 2nd day of culture. The hAECs were arranged in a typical cobblestone-like monolayer on the 4th and 7th day of culture. The primary hAECs highly expressed CD73, CD90, and CD29 of mesenchymal stem cell related surface markers, and were with no or low expressions of CD34 and HLA-DR of hematopoietic stem cell related surface markers. After 3 h of culture, hAEC-Exos were successfully endocytosed by HaCaT into the cytoplasm and gathered around the nucleus. After 12, 24, 36, 48, and 60 h of culture, the proliferation activity of HaCaT in hAEC-Exos group was significantly higher than that in PBS group (t=3.691, 10.861, 12.121, 10.531, 14.931, P<0.01). At 24, 48, and 72 h after scratch, the scratch healing rates of HaCaT in PBS group were significantly lower than those in hAEC-Exos group (t=3.342, 6.427, 5.485, P<0.05 or P<0.01). After 48 h of culture, the number of transmembrane HaCaT in hAEC-Exos group was significantly more than that in PBS group (t=5.385, P<0.01). After 24 h of culture, the protein expressions of p-mTOR and p-Akt in HaCaT of hAEC-Exos group were significantly higher than those in PBS group (t=4.240, 5.586, P<0.01), while the protein expressions of mTOR and Akt in HaCaT of the two groups were similar (P>0.05). After 24 h of culture, the protein expressions of p-mTOR and p-Akt in HaCaT of DMSO+hAEC-Exos group were significantly higher than those in DMSO+PBS group (t=6.155, 8.338, P<0.01) and LY294002+hAEC-Exos group (t=5.030, 3.960, P<0.01), while the protein expressions of mTOR and Akt in HaCaT of the three groups were similar (P>0.05). The proliferation activity of HaCaT in DMSO+hAEC-Exos group at 12, 24, 36, 48, and 60 h of culture was 0.78±0.05, 1.23±0.07, 1.60±0.09, 1.86±0.09, and 2.03±0.08, which was significantly higher than 0.46±0.04, 0.69±0.07, 0.98±0.08, 1.16±0.08, and 1.26±0.11 in DMSO+PBS group (t=4.376, 7.398, 8.488, 9.766, 10.730, P<0.01). The proliferation activity of HaCaT in DMSO+hAEC-Exos group at 24, 36, 48, and 60 h was significantly higher than 0.96±0.09, 1.20±0.08, 1.39±0.08, and 1.55±0.10 in LY294002+hAEC-Exos group (t=3.639, 5.447, 6.605, 6.693, P<0.05 or P<0.01). The scratch healing rates of HaCaT in DMSO+hAEC-Exos group at 24, 48, and 72 h after scratch were significantly higher than those in DMSO+PBS group (t=4.003, 6.349, 7.714, P<0.01) and LY294002+hAEC-Exos group (t=3.805, 4.676, 4.067, P<0.05 or P<0.01). After 48 h of culture, the number of transmembrane HaCaT in DMSO+hAEC-Exos group was significantly more than that in DMSO+PBS group and LY294002+hAEC-Exos group, respectively (t=7.464, 1.232, P<0.01). Conclusions: PI3K-Akt-mTOR pathway can promote the proliferation and migration of HaCaT in high glucose environment by mediating hAEC-Exos.

[Clinical features of 123 patients with hyperinsulinemic hypoglycemia auxiliarily diagnosed by 18F-DOPA-PET CT scanning].

Objective: To summarize the clinical features and therapeutic outcomes of patients with hyperinsulinemic hypoglycemia (HH) auxiliarily diagnosed by 18F-DOPA positron emission tomography (PET) CT scanning. Methods: The clinical data of 123 patients who were diagnosed with hyperinsulinemic hypoglycemia by comprehensive clinical diagnostic procedures in the Department of Pediatric Endocrinology and Inherited Metabolic Diseases, Children's Hospital of Fudan University between January 2016 and December 2020 were retrospectively analyzed. Clinical data such as gender, age of onset, province, concurrent serum insulin level measured during hypoglycemia, lesion type of pancreas by 18F-DOPA-PET CT scanning, genetic test results, and treatment were collected successively. The clinical features and therapeutic outcomes were compared between patients with focal and diffuse pancreatic lesions. T test, Rank sum test, and χ² test were used for comparison between groups. Results: A total of 123 patients with hyperinsulinemic hypoglycemia (72 males and 51 females), whose average age of onset was 3 days (ranging from 1 day to 4 860 days), were recruited from 24 provinces. The concurrent serum insulin level was 7.1 (0.4-303.0) mU/L during hypoglycemia. 18F-DOPA-PET CT scanning identified focal lesions in 25.2% (31/123) and diffuse lesions in 74.8% (92/123) of the patients; 64.2% (79/123) of the HH cases were found to have pathogenic gene variants, in which 88.6% (70/79) were found to have KATP channel related genes (61 in ABCC8 and 9 in KCNJ11 mutations). Thirty-seven patients (17 focal and 20 diffuse) received surgical treatment with a success rate of 67.6% (25/37). The effective rate of diazoxide for children with diffuse type was significantly higher than that of children with focal group (28.3% (26/92) vs. 9.7% (3/31), χ²=10.31, P=0.001). Conclusions: 18F-DOPA-PET CT scan can improve the success rate of surgery. Comprehensive diagnosis of the etiology of hyperinsulinemic hypoglycemia by genetic analysis and 18F-DOPA-PET CT scanning can result in better treatment and prognosis.

[Value of blood routine indexes and their ratios in judging the prognosis of adult patients with extensive burns].

Objective: To investigate the value of neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), and blood platelet count (BPC) in judging the prognosis of adult patients with extensive burns. Methods: From January 2012 to December 2018, 99 adult patients with extensive burns who met the inclusion criteria were admitted to Union Hospital of Fujian Medical University, including 76 males and 23 females, aged 18 to 75 (43±13) years. According to the prognosis, the patients were divided into survival group of 79 cases and death group of 20 cases. Their clinical data were retrospectively analyzed by the method of case-control study. The gender, age, total burn area, inhalation injury, use of mechanical ventilation and white blood cell count, neutrophil count, lymphocyte count, and BPC on post injury day (PID) 1, 3, and 7 were collected, and the NLR, PLR, difference value of BPC on PID 3 and PID 1 (ΔBPC3), difference value of NLR on PID 3 and PID 1 (ΔNLR3), difference value of PLR on PID 3 and PID 1 (ΔPLR3), difference value of BPC on PID 7 and PID 1 (ΔBPC7), difference value of NLR on PID 7 and PID 1 (ΔNLR7), difference value of PLR on PID 7 and PID 1 (ΔPLR7) of patients in the two groups were calculated. Data were statistically analyzed with Mann-Whitney U test, independent sample t test, chi-square test to screen the death-related factors of patients. Binary classification single factor and multifactor logistic regression analysis were used to analyze the death-related factors of patients. The receiver's operating characteristic (ROC) curve of the independent risk factor of death of patients predicting the prognosis of adult patients with extensive burns was drawn, and the area under the curve, the optimal threshold and its sensitivity and specificity were calculated. Results: (1) There were statistically significant differences in total burn area and use of mechanical ventilation of patients between the two groups (Z=-2.615, χ(2)=7.282, P<0.01). (2) On PID 1, there was statistically significant difference in NLR of patients between the two groups (Z=-2.414, P<0.05). On PID 3, there were statistically significant differences in BPC and ΔNLR3 of patients between the two groups (Z=-2.048, -2.780, P<0.05 or P<0.01). On PID 7, there were statistically significant differences in lymphocyte count, BPC, NLR, and ΔNLR7 of patients between the two groups (Z=-2.248, -2.231, -2.641, -3.669, P<0.05 or P<0.01). (3) Binary classification single factor logistic regression analysis showed that the total burn area, mechanical ventilation, BPC and NLR on PID 7, and ΔNLR7 were related to death of patients (odds ratio=1.038, 0.193, 0.990, 1.086, 1.105, 95% confidence interval=1.010-1.067, 0.062-0.598, 0.982-0.998, 1.012-1.165, 1.037-1.178, P<0.05 or P<0.01). Binary classification multifactor logistic regression analysis showed that ΔNLR7 was the independent risk factor of death of adult patients with extensive burns (odds ratio=1.090, 95% confidence interval=1.008-1.178, P<0.05). (4) The optimal threshold of ROC curve of ΔNLR7 for predicting the prognostic death of 97 adult patients with extensive burns was -0.073 4. The sensitivity under the optimal threshold was 65.0%, and the specificity was 78.5%. The area under the ROC curve was 0.776 (95% confidence interval=0.650-0.882, P<0.01). Conclusions: Dynamic monitoring of NLR and BPC is of great significance to assist in judging the prognosis of adult patients with extensive burns. ΔNLR7 is an independent predictor of death in adult patients with extensive burns, while PLR can not predict the death of adult patients with extensive burns.

[The expression of SnoN in human hypertrophic scar fibroblasts and the mechanism of its participation in hypertrophic scar formation].

Objective: To investigate the expression of SnoN in human hypertrophic scar fibroblasts and the mechanism of its participation in hypertrophic scar formation. Methods: Eight patients with hypertrophic scar after burn in need of surgery were admitted in our unit from January to October 2013, and then hypertrophic scar tissue and normal skin tissue of full-thickness skin donor site resected by surgery of the patients were collected. Hypertrophic scar fibroblasts and normal skin fibroblasts of patients were isolated with method of explant culture and then sub-cultured. Cells of the third to fifth passage were used in the following experiments. (1) The protein expressions of SnoN of hypertrophic scar fibroblasts and normal skin fibroblasts were assessed with Western blotting. (2) The mRNA expressions of SnoN of another batch of hypertrophic scar fibroblasts and normal skin fibroblasts were determined with reverse transcription polymerase chain reaction. (3) Another batch of hypertrophic scar fibroblasts and normal skin fibroblasts were treated with 10 ng/mL transforming growth factor beta1 (TGF-ß(1)) for 30 min, 1 h, 2 h, and 6 h, respectively, and then the protein expressions and mRNA expressions of SnoN of untreated cells and treated cells were detected as above. Data were processed with one way analysis of variance and independent sample t test. Results: (1) The protein expression of SnoN of hypertrophic scar fibroblasts was 0.020±0.003, significantly lower than that of normal skin fibroblasts (0.032±0.005, t=7.19, P<0.05). (2) The mRNA expression of SnoN of hypertrophic scar fibroblasts was 0.407±0.157, with no significant difference from that of normal skin fibroblasts (0.339±0.095, t=-1.29, P>0.05). (3) The protein expression of SnoN of normal skin fibroblasts was increased in a time-dependent fashion with the TGF-ß(1) stimulation, and the protein expressions of SnoN of cells treated with TGF-ß(1) for 30 min, 1 h, 2 h, and 6 h were significantly higher than those of untreated cells (with t values from 2.27 to 27.89, P values below 0.05). The protein expression of SnoN of hypertrophic scar fibroblasts was decreased in a time-dependent fashion with the TGF-ß(1) stimulation, and the protein expressions of SnoN of cells treated with TGF-ß(1) for 30 min, 1 h, 2 h, and 6 h were obviously lower than those of untreated cells (with t values from 10.80 to 13.85, P values below 0.05). (4) The mRNA expressions of SnoN of normal skin fibroblasts and hypertrophic scar fibroblasts were both increased in a time-dependent fashion with the TGF-ß(1) stimulation, and the mRNA expressions of SnoN of the two types of cells treated with TGF-ß(1) for 30 min, 1 h, 2 h, and 6 h were both significantly higher than those of untreated cells (with t values from 18.16 to 58.22, P values below 0.05). Conclusions: The protein expression of SnoN in hypertrophic scar fibroblasts is reduced, which weakens its inhibitory effect on TGF-ß(1) signal, thus amplifying the TGF-ß(1) signal, and it may participate in the formation of hypertrophic scar.

[Effect of inserted angle on the stability of loaded microscrews].

Objective: To evaluate the effect of different insertion angles on the osseointegration of loaded microscrews in beagle jaws. Methods: Forty-eight microscrews were inserted at four different angles (30°, 50°, 70° and 90°) into the interradicular zones between the mandibular first molar and third premolar in twelve beagles and the microscrews had been loaded with a force of 2 N immediately for 8 weeks. After microscrew-bone specimens fixed, the maximum output value (Fmax) of pull-out test was recorded and the histomorphological changes of hard tissue were observed. The bone-implant contact (BIC%) was quantitatively analyzed and the osseointegration of microscrew-bone interface was comprehensively evaluated. Results: Both Fmax and BIC% values of microscrews were influenced by the insertion angles. The maximum value of Fmax was (385±23) N in the group with 50° angle, and the minimum value was (198±16) N in the group with 30° angle(P <0.05). The maximum value of BIC% was (59.1±6.0)% in the group with 70° angle, and the minimum value was (30.2±3.2)% in the group with 30° angle (P <0.05). Histomorphology observation revealed that in peri-screws region, the various degree of bone remodeling was found in different angle samples. Conclusions: The insertion angles (50°and 70°) were favorable to the stability of the microscrew.

Distribution and localization of abnormally expressed OPTN proteins in RGC5 retinal ganglion cells and their effects on subcellular morphology.

The objectives of this study were to investigate the distributions of abnormally expressed optineurin (OPTN) proteins in retinal ganglion cells (RGC5s) of transgenic rats and their effects on subcellular morphological structures. Green fluorescent protein labeled EGFP wild-type (OPTN(WT)), E50K mutant type (OPTN(E50K)), and OPTN siRNA (si-OPTN) eukaryotic expression plasmids were constructed and transfected into RGC5s. Intracellular structures were labeled with organelle specific fluorescent dyes. Construct localization and cell morphologies were visualized by confocal fluorescence microscopy. OPTN(WT) was observed to be distributed as fine punctate fluorescent particles in the cytoplasm around the nucleus, along with exhibiting nuclear expression. OPTNE50K exhibited similar distribution but with non-uniform fluorescence particle size. si-OPTN distribution was similar to that of EGFP: uniform across the cytoplasm and nucleus. Compared with the negative control group, OPTN(WT), and OPTN(E50K) and to a lesser degree pEGFP-transfected cells exhibited fracture and loss of myofilament proteins and mitochondrial swelling and cytoplasmic accumulation, along with abnormal lysosomal distribution and increased volume, and Golgi fragmentation. However, si- OPTN transfected cells exhibited no significant damage. Therefore, we demonstrated that the E50K mutation disrupts the uniformity of OPTN protein distribution upon exogenous overexpression. Furthermore, these results suggested that si-OPTN transfection, and thus potentially OPTN knockdown, did not impact subcellular morphology of RGC5 cells, whereas transfection, especially when combined with wild-type or mutant OPTN expression, led to substantial abnormalities in subcellular morphological structures. These findings lay a foundation for further research into the function of the OPTN protein.

Effects of abnormal optineurin expression on the survival of the rat retinal ganglion cell line RGC-5.

The OPTN gene is thought to be associated with certain types of glaucoma and the function of the protein for which it encodes, optineurin, has been extensively researched, but with contradictory results. We explored the effects of abnormal optineurin expression on the survival of the rat retinal ganglion cell line RGC-5. Plasmids expressing wild-type (WT) or E50K mutant optineurin, or OPTN-specific double-hairpin small interfering RNA (si-RNA), were transfected into RGC-5 cells. The effects on cell survival were monitored by observation of cell morphology and propidium iodide and Hoechst 33342 fluorescent staining, while expression of optineurin was visualized by fluorescence microscopy. Abnormal optineurin expression influenced the survival of RGCs in vitro, as apoptosis was induced by increased WT and E50K mutant optineurin, while a reduction in apoptosis was observed in cells transfected with OPTN-siRNA. Similar results were also observed in transfected cells treated with apoptotic stimuli. Overexpression of WT and mutant E50K protein resulted in greater cell death, while downregulation decreased RGC-5 apoptosis.

Well-balanced commensal microbiota contributes to anti-cancer response in a lung cancer mouse model.

The intestinal microflora affects inflammation and immunity, not only locally at the mucosal level but also systemically, raising the question of whether the microflora affects inflammatory processes that contribute to cancer and its therapy. Prebiotics have also been found to play an antitumor role that is not limited to the gut. We investigated the antitumor roles of the intestinal microbiota using the Lewis lung cancer mouse model. In mice treated with cisplatin combined with ABX (an antibiotic cocktail of vancomycin, ampicillin, and neomycin), which can destroy the host commensal microflora, the tumor size was larger than in mice on a single treatment of cisplatin. Moreover, the survival rate of mice treated with cisplatin combined with ABX was significantly reduced. In contrast, mice treated with cisplatin combined with Lactobacillus bacteria had smaller tumors and an improved survival rate. Further study on gene expression indicated that ABX can partially impair the function of cisplatin by upregulating the expression of VEGFA and downregulating the expression of BAX and CDKN1B. The expression of IFN-γ, GZMB, and PRF1 in the CD8(+) T cells of these mice was reduced by ABX, indicating an immuno-enhancement role of commensal microbiota. Conversely, Lactobacillus co-treatment mice showed an enhanced antitumor response with upregulated IFN-γ, GZMB, and PRF1 expression. We conclude that the commensal microbiota contributes to the anti-lung cancer response and probiotics co-treatment can enhance the antigrowth and proapoptotic effects of cisplatin.

Quantitative evaluation of heavy metals in solid residues from sub- and super-critical water gasification of sewage sludge.

Solid residues (SRs) are important byproducts of sub- and super-critical water gasification of sewage sludge (SS). In this study, the quantitative evaluation of heavy metals (HMs) in SRs, compared with SS, is applied in terms of potential ecological risks, pollution levels, and both bioavailability and eco-toxicity. The results show the bioavailability and eco-toxicity of HMs in SRs decrease, although the total concentration of HMs increased, particularly in the bioavailable fraction of Cu, which decreased nearly 97%. The geo-accumulation and potential ecological risk index indicated that the gasification process increased contamination by two levels (to the maximum), while the overall risk was in keeping with SS. However, based on the risk assessment code, each tested HM exhibited lower environmental risk after gasification, especially for Cd, which drastically dropped from 66.67 (very high risk) in SS to 0.71 (no risk) in SRs, with a reaction temperature of 375°C for 60 min.

Partial oxidative gasification of municipal sludge in subcritical and supercritical water.

Subcritical and supercritical water gasification of dewatered sewage sludge obtained from a typical municipal wastewater treatment plant was investigated in a one-litre high-pressure autoclave at temperatures of 300-400 degrees C and pressures of 17.5-23.5 MPa. The sludge (without catalyst) was gasified at subcritical and supercritical water conditions, with different reaction times ranging from 30 to 60 min. The results showed that gaseous product yield increased with increasing temperature and reaction time. Gas products consisted primarily of hydrogen, carbon dioxide, methane, carbon monoxide and other light hydrocarbons. The liquid products contained high levels of organic matter, ammonia nitrogen and a few heavy metals. Compared with the landfilling of sewage sludge, the solid residues were in accordance with the Chinese standard for sludge quality in co-landfilling even without further treatment. In addition, the heavy metals in solid products exhibited more stable characteristics attributable to the reduced leaching toxicity after supercritical water gasification.

Changes in composition a metabolism of caecal microbiota in rats fed diets supplemented with copper-loaded chitosan nanoparticles.

The study was conducted to investigate the effect of copper-loaded chitosan nanoparticles on the composition and metabolism of the caecal microbiota in rats. Forty male Sprague-Dawley (SD) rats (average body weight of 82 ± 5 g) were randomly assigned to five groups (n = 8). The dietary treatments were: (i) basal diet, (ii) basal diet + 80 mg/kg BW CuSO(4), (iii) basal diet + 80 mg/kg BW chitosan (CS-I), (iv) basal diet + 80 mg/kg BW copper-loaded chitosan nanoparticles (CSN-I) and (v) basal diet + 160 mg/kg BW copper-loaded chitosan nanoparticles (CSN-II). The trial lasted 21 days. The results showed that compared with control, Average day gain (ADG) of group CSN-I and CSN-II increased (p < 0.05). No significant difference was found in CuSO(4) or CS-I-treated groups (p > 0.05). There were no effects of these treatments on average day feed intake (ADFI) of rats (p > 0.05). The counts of Bifidobacteria and Lactobacillus in group CSN-II were higher than those of the control group (p < 0.05), while the counts of total aerobes, total anaerobes, Salmonella, Clostridium and coliform were lower than those of the control (p < 0.05). The activity of ß-glucuronidase in group CSN-I or CSN-II was significantly depressed (p < 0.05), while the activities of α-galactosidase and ß-galactosidase were enhanced significantly (p < 0.05). The pH of the caecum digesta and the concentration of propionate and butyrate in group CSN-I and CSN-II were higher than those of the control group (p < 0.05). No significant difference was found in these parameters in CuSO(4) or CS-I-treated groups (p > 0.05). The results indicate that the microbiota and environment of caecum are beneficially changed by the administration of copper-loaded CSN.

Effect of chromium nanocomposite supplementation on growth hormone pulsatile secretion and mRNA expression in finishing pigs.

The study was conducted to investigate the effect of chromium nanocomposite (CrNano) on growth hormone (GH) pulsatile secretion and pituitary GH mRNA expression in finishing pigs. Fifty-four crossbred pigs (65.57 ± 1.05 kg initial weight) were randomly allotted to one of the three treatments, with three replicate pens per treatment and six pigs per pen. Pigs were offered one of the four diets including a control diet or a control diet supplemented with 200 µg/kg chromium from either chromium chloride (CrCl3) or CrNano for 35 days. During the trial, all pigs were given free access to feed and water. After completion of the feeding trial, blood samples were taken via auriculares at 15 min intervals for 3 h and three pigs from each treatment were slaughtered to collect pituitary to determine GH mRNA level. The results of GH dynamic secretion showed that supplemental CrNano increased the mean level, the lowest value, peak value and peak duration of GH significantly, while there was no significant effect on peak amplitude. Pituitary mRNA expression of GH was improved significantly in pigs fed diet containing Cr from CrNano. These results indicated that CrNano increased GH mRNA expression and secretion in finishing pigs.

Effects of beta-glucanase and xylanase supplementation on gastrointestinal digestive enzyme activities of weaned piglets fed a barley-based diet.

The effects of supplementing a barley-based diet for weaned piglets withexogenous beta-glucanase and xylanase on gastrointestinal digestiveenzyme activities were investigated. Thirty-six cross-bred weaned pigletswere randomly assigned to two groups with three pens based on sexand mass. Each group was fed on the diet based on barley with or withoutadded beta-glucanase and xylanase (0.15%) for a 4-week period. Theresults showed that enzyme supplementation improved growth performanceof piglets significantly (p < 0.05), but had no effect (p = 0.091)on average daily feed intake. The results also showed that supplementationof beta-glucanase and xylanase had no effect on pepsin activity in gastriccontents but slightly decreased (p = 0.092) the pepsin activity ingastric mucosa. Meanwhile, no effect of enzyme supplementation ontrypsin activity in duodenal contents was observed. However, the activitiesof amylase and lipase in duodenal contents were significantly(p < 0.05) decreased, whereas the activities of maltase, sucrase andgamma-glutamyl transpeptidase (gamma-GT) in jejunal and ileal mucosa wereenhanced significantly (p < 0.05). The improvement of disaccharidaseand gamma-GT activity may be attributed to the positive impacts of exogenousenzymes on digestion and absorption of the nutrients. In conclusion,the current results indicated that supplementation with enzymes in barley-based diets could improve the growth performance of piglets,decrease the activities of amylase and lipase in duodenal contents andincrease the activities of disaccharidase and gamma-GT in jejunal and ilealmucosa.

Developmental changes of carcass composition, meat quality and organs in the Jinhua pig and Landrace.

The present study was aimed to compare the developmental changes of carcass composition, meat quality characteristics and organ weight in pigs of different breeds. Six pigs (sex balance) of each breed were slaughtered at 35, 80 and 125 days of age, respectively. The carcass was chilled and the left carcass side was dissected into bone, lean meat, fat and skin; additionally, organ weight and meat quality parameters were observed. Carcasses of the Jinhua pig were lighter (P < 0.001), contained less lean meat percentage (P < 0.01) and more carcass fat percentage (P < 0.05) than did carcasses of the Landrace. L*-values were lower in Jinhua pigs than in Landrace at 125 days of age (P < 0.05), but the Jinhua pig had higher a*-values compared with Landrace at the age of 80 days (P < 0.01) and 125 days (P < 0.01), respectively. In addition, Jinhua pigs showed lower colour scores (P < 0.05), higher intramuscular fat (IMF) percentage (P < 0.05), less marbling scores (P < 0.05) and lower drip loss (P < 0.05) than Landrace. For organ weight, Jinhua pigs had higher relative heart weight at the age of 80 days (P < 0.05) and 125 days (P < 0.001), and higher relative liver weight at 125 days of age (P < 0.01) than that of Landrace. In addition, the relative kidney weight was heavier (P < 0.001) in the Jinhua pig than in the Landrace during the whole experiment. These results indicated that developmental changes of carcass composition, meat quality parameters and organ weight displayed breed differences. Jinhua pigs were fatter than Landrace but the former had better quality characteristics in the meat.

Oral vaccination with envelope protein VP28 against white spot syndrome virus in Procambarus clarkii using Bacillus subtilis as delivery vehicles.

AIMS: To achieve high-level expression and secretion of active VP28 directed by a processing-efficient signal peptide in Bacillus subtilis WB600 and exploit the possibility of obtaining an oral vaccine against white spot syndrome virus (WSSV) using vegetative cells or spores as delivery vehicles. METHODS AND RESULTS: The polymerase chain reaction (PCR)-amplified vp28 gene was inserted into a shuttle expression vector with a novel signal peptide sequence. After electro-transformation, time-courses for recombinant VP28 (rVP28) secretion level in B. subtilis WB600 were analysed. Crayfish were divided into three groups subsequently challenged by 7-h immersion at different time points after vaccination. Subgroups including 20 inter-moult crayfish with an average weight of 15 g in triplicate were vaccinated by feeding coated food pellets with vegetative cells or spores for 20 days. Vaccination trials showed that rVP28 by spore delivery induced a higher resistance than using vegetative cells. Challenged at 14 days postvaccination, the relative per cent survival (RPS) values of groups of rVP28-bv and rVP28-bs was 51.7% and 78.3%, respectively. CONCLUSIONS: The recombinant B. subtilis strain with the ability of high-level secretion of rVP28 can evoke protection of crayfish against WSSV by oral delivery. SIGNIFICANCE AND IMPACT OF THE STUDY: Oral vaccination by the B. subtilis vehicle containing VP28 opens a new way for designing practical vaccines to control WSSV.

Chromium nanoparticle exhibits higher absorption efficiency than chromium picolinate and chromium chloride in Caco-2 cell monolayers.

This study was conducted to determine whether chromium nanoparticle (CrNano) exhibited higher absorption efficiency and possessed unique absorption mechanism in comparison to chromium picolinate (CrPic) and chromium chloride (CrCl(3)), as was postulated by previous reports. Twenty-one-day-old Caco-2 cell monolayers grown on semipermeable membranes in Snapwell tissue culture bichambers were incubated with CrNano, CrPic or CrCl(3) to examine their transport and uptake respectively. In the concentration range of 0.2-20 micromol/l, transport of CrNano, CrPic and CrCl(3) across Caco-2 monolayers both in apical-to-basolateral and basolateral-to-apical direction was concentration-, and time-dependent, and temperature independent. The apparent permeability coefficient (P(app)) of CrNano was between 5.89 and 7.92 x 10(-6) cm/s and that of CrPic and CrCl(3) was between 3.52 and 5.31 x 10(-6) cm/s and between 0.97 and 1.37 x 10(-6) cm/s respectively. Uptake of CrNano, CrPic and CrCl(3) by both apical and basolateral membranes was concentration- and time-dependent. Uptake of CrNano by apical membrane was significantly (p < 0.05) decreased when the incubation temperature was reduced from 37 degrees C to 4 degrees C. The transport efficiency of CrNano, CrPic and CrCl(3) after incubation for 120 min at 37 degrees C was 15.83% +/- 0.76%, 9.08% +/- 0.25% and 2.11% +/- 0.53% respectively. The uptake efficiency of CrNano, CrPic and CrCl(3) was 10.08% +/- 0.76%, 4.73% +/- 0.60% and 0.88% +/- 0.08% respectively. It was concluded that the epithelial transport of CrNano, CrPic and CrCl(3) across the Caco-2 cell monolayers was mainly via passive transport pathways. In addition, CrNano exhibited considerably higher absorption efficiency than both CrPic and CrCl(3) in Caco-2 cell monolayers.

Oral allergy syndrome and anaphylactic reactions in BALB/c mice caused by soybean glycinin and beta-conglycinin.

BACKGROUND: Soybean protein is used in a number of food products but is also a common cause of food allergy. Soybean glycinin and beta-conglycinin represent up to one-third of protein in the soybean. Many reports have indicated that glycinin and beta-conglycinin have been characterized as major soybean allergens involved in food hypersensitivity. OBJECTIVE: To investigate oral allergy syndrome and anaphylactic reactions in BALB/c mice caused by soybean glycinin and beta-conglycinin with an intragastric feeding protocol without using an adjuvant. METHODS: BALB/c mice were sensitized by gavages with glycinin and beta-conglycinin, and allergen-specific IgE and IgG1 responses were studied by a passive cutaneous anaphylaxis assay. Serum histamine release and blood pressure were measured according to other methods. Epithelium and mast cell dye used the method of light microscopy. RESULTS: Sensitization with soybean allergens induced high levels of antigen-specific IgE and IgG1 and increased serum histamine in BALB/c mice. Percentiles of intact mast cell of small intestine in mice sensitized with glycinin and beta-conglyinin significantly decreased for 28 days. Degranulation of mast cells and damage of the epithelium in the small intestine of mice sensitized with globulins were observed. The level of blood pressure in sensitized mice reached a minimum at 3 h. CONCLUSION: Soybean-specific IgE and IgG1 antibodies increased, with high levels of histamine release, severe degranulation of mast cells and damage of the epithelium of small intestine in mice sensitized with glycinin and beta-conglyinin.

Efficacy of chromium(III) supplementation on growth, body composition, serum parameters, and tissue chromium in rats.

Chromium(III) is often claimed to have a positive effect on body composition, while the responses in researches with supplementation of different chemical form of chromium are various and inconsistent. We have studied the effects of 6 weeks of treatment with three different forms of chromium (300 mug/kg) as chromium chloride, chromium tripicolinate, and chromium nanocomposite (CrNano) on growth, body composition, serum parameters, and tissue chromium in rats. The supplementation of CrNano significantly increased average daily gain, food efficiency, and lean body mass and decreased fat mass and body fat proportion and serum levels of glucose, urea nitrogen, triglyceride, and insulin. Chromium contents in liver, kidney, and hind leg muscle were increased significantly with the addition of CrNano in diet. The results indicate that chromium nanocomposite has higher efficacy on growth and body composition compared to the traditional chromium agents.

Effect of supplementation of green tea polyphenols on the developmental competence of bovine oocytes in vitro.

The objective of the present study was to examine the effect of green tea polyphenols (GTPs) supplementation during in vitro maturation, in vitro fertilization, and in vitro culture on the developmental competence of bovine oocytes. Cumulus-oocyte complexes aspirated from the ovaries were matured in vitro (38.5 degrees C for 24 h) and fertilized (38.5 degrees C for 15-18 h) and embryos were cultured (38.5 degrees C for 192 h) in a defined conditioned medium with or without GTPs supplementation. The GTPs used in the present study contained 99% catechin derivatives, with the major components being 50% (-)-epigallocatechin gallate, 22% (-)-epicatechin gallate, 18% (-)-epigallocatechin, and 10% (-)-epicatechin. Four replicate trials were done for each type of experiment. GTPs supplementation (15 microM) of the maturation medium led to a significant increase in the rate of blastocyst formation (34.0 vs 21.4%, P < 0.05). However, the rate of blastocyst formation was not improved when higher GTPs concentrations (20 or 25 microM) were added to the in vitro maturation medium. During in vitro fertilization, supplementation with higher GTPs concentrations (20 or 25 microM) significantly reduced the rate of blastocyst formation (P < 0.05). Supplementation of the culture medium with 15 microM GTPs improved the rate of blastocyst formation, while higher GTPs concentrations (25 microM) significantly reduced embryo development (P < 0.05). In conclusion, these results demonstrate that supplementation with GTPs at low concentration (15 microM) during in vitro maturation and in vitro culture improved the developmental competence of bovine oocytes.