Objective To construct gene silence adenovirus vector targeting both transglutaminase 2 （TG2） and Mer receptor tyrosine kinase （Mertk） synchronously and detect the gene silence function of it. Methods The interfering plasmids targeting TG2 protein and Mertk protein were constructed firstly， then the H1 promoter and RNA interfering （RNAi） sequence were cut and ligated to pAdTrack for constructing pAdTrack/TG2/Mertk. The pAdTrack/TG2/Mertk was transfected into BJ5183 bacterial cells which contained pAdEasy-1， then the plasmid was detected by enzyme digestion after recovery. Adenovirus were harvested after that pAdTrack/TG2/Mertk was infected into HEK293 cells. The virus titer was measured after repeated amplification. The RAW264.7 cells were infected by pAdTrack/TG2/Mertk， pAdTrack/TG2， pAdTrack/Mertk， and pAdTrack/green fluorescent protein （GFP）， respectively. Then the expression levels of TG2 protein and Mertk protein of mouse macrophages were detected by Western blot after infection. Results The virus titer of pAdTrack/TG2/Mertk plasmid was 6.13×1010GFU/mL. The pAdTrack/TG2/Mertk plasmid which contained 2 promoters and 2 RNAi sequences was identified successfully by enzyme digestion. Compared with pAdTrack/GFP group and pAdTrack/Mertk group （there was no significant differece between the 2 groups）， the expression levels of TG2 protein of mouse macrophages which infected with pAdTrack/TG2/Mertk or pAdTrack/TG2 decreased obviously （P＜0.01）， but there was no significant difference between the later 2 groups. Compared with pAdTrack/GFP group and pAdTrack/TG2 group （there was no significant difference between the 2 groups）， the expression levels of Mertk protein of mouse macrophages which infected with pAdTrack/TG2/Mertk or pAdTrack/Mertk decreased obviously too （P＜0.01）， but there was no significant difference between the later 2 groups. Conclusion Gene silence adenovirus vector plasmid targeting both TG2 and Mertk synchronously is constructed successfully， and the pAdTrack/TG2/Mertk can reduce the expressions of TG2 protein and Mertk protein of mouse macrophages obviously.
Objective To investigate the effect of mesenteric lymphatic duct liagtion and glutamine enteral nutrition on intestine and distant organs in intestinal ischemia/reperfusion injury. Methods Forty male SD rats undergoing gastrostomy were randomly assigned into 5 groups (n=8): sham operation group, normal enteral nutrition group, normal enteral nutrition+lymphatic duct ligation group, glutamine group and glutamine+lymphatic duct ligation group. Sham operation group only received laparotomy after 7 days of full diet, the other four groups were subjected to 60 min of intestinal ischemia after 7 days of enteral nutrition, and the two lymphatic duct ligation groups were plus mesenteric lymphatic duct ligation. The original nutrition continued 3 days after reperfusion. Intestinal permeability was detected on day 1 before reperfusion, day 1 and 3 after reperfusion. Intestinal morphology was observed, endotoxin, D-lactate and diamine oxidase levels in serum, and apoptotic index in lung tissue were detected on day 3 after reperfusion. Results The intestinal permeability in each group was significantly increased on day 1 after reperfusion (Plt;0.05), and which in normal enteral nutrition+lymphatic duct ligation group and glutamine+lymphatic duct ligation group were significantly decreased on day 3 after reperfusion (Plt;0.05). The mucosal thickness and villus height of ileum and mucosal thickness of jejunium in glutamine+lymphatic duct ligation group were significantly higher than those in other groups (Plt;0.05), and villus height of ileum in glutamine group was higher than that in normal enteral nutrition group (Plt;0.05); those morphology indexes in normal enteral nutrition+lymphatic duct ligation group were higher than those in normal enteral nutrition group, but there was no statistical signification (Pgt;0.05). Apoptosis index of lung tissue in lymphatic duct ligation groups was significant lower than that in no-ligation groups (Plt;0.05). Levels of endotoxin, D-lactate, and diamine oxidase in lymphatic duct ligation groups had downward trends compared with no-ligation groups, but there was no statistical signification (Pgt;0.05). Conclusions Intestinal ischemia/reperfusion injury of rats can cause intestinal permeability increase, bacterial endotoxin translocation and systemic inflammatory response. Mesenteric lymphatic duct ligation and glutamine enteral nutrition intervention can weak lung tissue damage, increase thickness of intestinal mucosa, maintain intestinal barrier function, reduce endotoxin translocation and attenuate systemic inflammatory response. Enteral nutrition with glutamine was better than normal enteral nutrition.
ObjectiveTo evaluate the efficacy of intravenous glutamine on patients with severe acute pancreatitis. MethodsThe Cochrane Library, PubMed, EMbase, CNKI and CBM databases were searched up to January 2013. Randomized controlled trials (RCTs) that compared non-glutamine nutrition with intravenous glutamine supplemented nutrition in patients with severe acute pancreatitis were included. A method recommended by the Cochrane Collaboration was used to perform a meta-analysis of those RCTs. ResultsFour RCTs involving a total of 190 participants were included. Analysis of these RCTs revealed the presence of statistical homogeneity among them. Results showed that glutamine dipeptide had a positive effect on reducing the mortality rate[OR=0.26, 95%CI (0.09, 0.73), P=0.01], length of hospital stay[WMD=-4.85 d, 95%CI (-6.67, -3.03) d, P<0.001], and the rate of complications[OR=0.41, 95%CI (0.22, 0.78), P=0.006]. No serious adverse effects were found. ConclusionCurrent best evidence demonstrates that glutamine is effective for severe acute pancreatitis. Further high quality trials are required and parameters of nutritional condition and hospital cost should be considered in future RCTs with sufficient size and rigorous design.
ObjectiveTo study the growth of adipose-derived stem cells (ADSCs) planted in three-dimensional (3D) materials, a 3D cultured ADSCs system based on microbial transglutaminase (mTG) enzyme crosslinked gelatin hydrogel was constructed. MethodsADSCs were isolated from the subcutaneous adipose tissue of a Sprague Dawley rat by collagenase digestion and centrifugation, and were cultured for passage. The mTG enzyme crosslinked gelatin hydrogel was firstly synthesized by mixing gelatin and mTG, and then the ADSCs were encapsulated in situ (2D environment) and cultured in the 3D materials (3D environment). The morphology and adhesion of cells were observed by inverted phase contrast microscope. In addition, HE staining and Masson staining were carried out to observe the distribution of cells in the material. Living and death situation of ADSCs in the materials was observed by fluorescence microscope and laser scanning confocal microscopy. Scanning electron microscopy was used to observe the adhesion of ADSCs on hydrogel surface. Alamar-Blue method was used to detect the proliferation of ADSCs in the hydrogel. Moreover, the results were compared between the cells cultured in 2D environment and those in 3D environment. ResultsThe result of 2D culture showed that ADSCs grew well on the hydrogel surface with normal functioning and had good adhesion. The results of 3D culture showed that ADSCs grew well in 3D cultured mTG enzyme crosslinked gelatin hydrogel, and presented 3D shape. Cells obviously extended in all directions. The number of apoptotic cells was very small. The cells of 3D culture at each time point was significantly less than that of the conventional culture cells, difference was statistically significant (P < 0.05). But after 8 days culture, the proliferation of the cells cultured in the mTG enzyme crosslinked gelatin hydrogel increased more quickly. ConclusionADSCs can grow well with good adhesion and show high viability in 3D culture system constructed by mTG enzyme crosslinked gelatin hydrogel.
Objective To study the effects of two different tidal volume mechanical ventilation on lipopolysaccharide( LPS) -induced acute lung injury( ALI) , and explore the effects of glutamine on ALI.Methods Thirty male Sprague-Dawley rats were randomly divided into three groups. After anesthesia and tracheotomy were performed, the rats were challenged with intratracheal LPS ( 5mg/kg) and received ventilation for 4 hours with small animal ventilator. Group A received conventional tidal volume, while groupB received large tidal volume. Group C received large tidal volume as well, with glutamine injected intravenously 1 hour before ventilation. Arterial blood gases were measured every one hour. 4 hours later, the rats were killed by carotid artery bleeding. The total lung wetweightwas measured and lung coefficient ( total lung wet weight /body weight ×100) was counted. WBCs and neutrophils in BALF were counted. Protein concentration, TNF-α, IL-6, and cytokine-induced neutrophil chemoattractant-1 ( CINC-1) levels in BALF,myeloperoxidase ( MPO) , and superoxide dismutase ( SOD) levels in the lung were assayed respectively.Results PaO2 and SOD levels decreased more significantly in group B than those of group A. The lung coefficient, WBCs, neutrophils, protein, TNF-α, IL-6, and CINC-1 levels in BALF, MPO levels in lung increased more significantly in group B than those of group A. PaO2 and SOD levels were significantly higher in group C than those of group B. The lung coefficient, WBCs, neutrophils, protein, TNF-α, IL-6, and CINC-1 levels in BALF,MPO levels in lung were significantly lower in group C than those of group B. Conclusion Large tidal volume mechanical ventilation aggravates LPS-induced ALI, and glutamine has obviouslyprotective effects.
ObjectiveTo summarize the current status of research in nutritional support for glutamine after hepatectomy.MethodThe literatures on nutritional support of glutamine after hepatectomy in recent years were reviewed by searching domestic and foreign literatures.ResultsThe administration of glutamine up-regulated the expression of liver regeneration genes after partial hepatectomy in malnourished rats, and then stimulated cell mitosis by paracrine and endocrine cells, affecting the uptake of amino acids by hepatocytes and intestinal cells, and promoting hepatocyte proliferation. In clinical applications, glutamine could improve postoperative liver function and immune function, reduce the incidence of infectious complications, then relatively shorten the length of hospital stay, and improve the clinical outcome of patients.ConclusionGlutamine is beneficial to the recovery of liver function and has clinical application value.
Objective To explore the effect and mechanism of glutamine to the aberrant crypt foci (ACF) in rat injured by acetic acid. Methods Thirty Wistar rats were averagely divided into three groups: control group, acetic acid group and glutamine group. The colon of the rat was infused with 1% acetic acid. Started to gavage with glutamate two days after modeling glutamine group. The injured colons were studied after fourteen days with light and scanning electronic microscope. Paraffin sections of specimens were prepared and stained with HE. The colon crypts were isolated by HCl digestion method. The expressions of CD44 and ICAM-1 in the epithelial cell of the large intestine mucosa were detected by immunohistochemistry method. Results On the days of 14， the number of ACF in the glutamine group were remarkably decreased as compared with that of the acetic acid group and a branch-like. The expressions of ICAM-1 and CD44 (every 1 000 cells) were 302.1±30.1 and 298.6±28.3 in glutamine group, 223.6±23.5 and 221.5±28.6 in control group, 198.5±19.5 and 215.3±17.8 in acetic acid group, respectively. While the expressions of CD44 and ICAM-1 in intestine were increased remarkably in the glutamine group compared with the control group and acetic acid group (P＜0.05). Conclusion Glutamine could decrease the formation of the ACF injured by acetic acid. Increasing the expressions of CD44 and ICAM-1 may be one of the important factors to decrease the ACF.
ObjectiveTo evaluate the effect of glutamineenhanced enteral nutritional support on elder patients after total gastrectomy. MethodsA total of eightyfour cases of elder patients receiving total gastrectomy were included in this study from February 2008 to August 2010. The patients were randomly divided into three groups: glutamineenhanced enteral nutrition (Gln) group, enteral nutrition (EN) group and parenteral nutrition (PN) group. The complications and hospital stay after operation were compared, and the levels of serum total protein, albumin, proalbumin， and transferrin of patients were measured before operation, on 2 d and 10 d after operation, respectively. Furthermore, the percentage of CD4 and CD8 T cells, CD4/CD8 ratio， and the levels of serum IgM and IgG of patients in peripheral blood before and after operation were detected. ResultsNutritional therapy was successfully performed in patients of three groups. The anal exhaust time and hospital stay after operation of patients in Gln group and EN group were significantly lower than those in PN group (Plt;0.05). The difference of postoperative complications and digestive tract symptoms of patients in three groups was not obvious (Plt;0.05). Anastomotic fistula occurred in one patient of PN group on 6 d after operation and was cured by conservative treatment for 54 d. The difference of total protein, albumin, proalbumin, and transferrin levels of patients in three groups before operation was not significant (Pgt;0.05), and these indexes fell dramatically on 2 d after operation and lower than before operation (Plt;0.05), although the intergroup difference was not statistically significant (Pgt;0.05). On 10 d after operation, all indexes recovered in different extent, while those data in Gln group and EN group were significantly higher than those on 2 d after operation (Plt;0.05). The levels of total protein, albumin, and proalbumin of patients in Gln group and EN group were markedly higher than those in PN group (Plt;0.05), although there was no difference between the former groups (Pgt;0.05). The difference of several immunological parameters of patients in three groups before operation was not significant (Pgt;0.05). On 10 d after operation, the percentage of CD4 and CD8 T cells, CD4/CD8 ratio, and the levels of serum IgM and IgG of patients in Gln group returned and even exceeded the preoperative results, which were significantly higher than those in EN group and PN group other than IgM (Plt;0.05). The postoperative results of all parameters except IgG in EN group were significantly lower than preoperative results in patients of EN group and PN group (Plt;0.05). ConclusionsIt is safe and feasible to elder patients who had received total gastrectomy and perioperative glutamine-enhanced nutritional support, which can improve nutrition and immune status, promote the recovery and reduce the duration of hospital stay, and nutritiional support after total gastrectomy is one of the optimal choices for these patients.