Involvement of PD-1+CD4+ T cells in the development of traumatic tracheal stenosis by regulating the IL-17/STAT3 pathway.

Studies have highlighted an upregulation of PD-1 expression in CD4+ T cells, which accelerates lung fibrosis by activating the IL-17/STAT3 pathway, leading to IL-17A and TGF-ß1 secretion. However, the relation with traumatic tracheal stenosis (TS) remains unexplored. Our analysis found significant increases in PD-1+CD4+ T cells, IL-17A, and TGF-ß1 in the TS patients (n = 10). The cellular model used CD4+ T cells co-cultured with bronchial fibroblasts while the animal model used a nylon brush to scrape the damaged tracheal mucosa. Interventions with PD-1 and STAT3 inhibitors both in vitro (n = 5) and in vivo (n = 6) showed decreased expression of TGF-ß1 and IL-17A in CD4+ T cells, decreased collagen I synthesis in vitro, and reduced tractal fibrosis in vivo. Furthermore, PD-1's modulation of the STAT3 was evident. This research unveils PD-1+CD4+ T cells' role in TS, thus suggesting a novel immunotherapeutic strategy to counteract tracheal fibrosis.

[Implementation and controversies of enhanced recovery after surgery in pancreaticoduodenectomy].

The concept of enhanced recovery after surgery(ERAS) has been widely accepted and applied in clinical practice.However,as one of the most complex surgical procedures in abdominal surgery,pancreaticoduodenectomy is characterized by long operation time,high incidence rate of postoperative complications and delayed recovery,there still remain some controversies about application of ERAS approaches in perioperative managements of pancreaticoduodenectomy.Although more and more studies has revealed the safety and efficacy of ERAS approaches in pancreaticoduodenectomy,the implementation of ERAS approaches should be still individualized in clinical practice to ensure safety of the patients.

[Effects of different drugs on bronchial stenosis by TGF-ß/mTOR signaling pathway in rabbit model].

Objective: To investigate the effect of different drugs on tracheal stenosis caused by transforming growth factor-ß/rapamycin target protein (TGF-ß/mTOR) signaling pathway. Methods: Thirty rabbits were randomly divided into normal control group, normal saline group, penicillin group, budesonide group and erythromycin group. The normal control group was not treated,and tracheal stenosis models were established in the other groups. From the 1st to 10th day after modeling, each group was respectively administered with normal saline (0.75 ml/kg, 2 times/d), intramuscular injection of penicillin (40 000 U/kg, 2 times/d), gastric administration of erythromycin (12.5 mg/kg, 2 times/d), inhalation of budesonide (0.05 mg/kg, 2 times/d). Rabbits were sacrificed on the 11th day after surgery, and tracheal specimens were collected to measure the degree of tracheal stenosis. Relative mRNA expression level of interleukin-6 (IL-6), transforming growth factor-ß (TGF-ß), Type Ⅰ collagen (COL-1), Type Ⅲ collagen (COL-3), and Sirtuin 1 (SIRT-1) were detected by Real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR); protein expression of mTOR, phosphorylated protein kinase B (p-AKT), vascular endothelial growth factor (VEGF),SIRT-1 were detected by immunohistochemical analysis; protein expression of nuclear factor κB (NF-κB),phosphorylated nuclear factor κB (p-NF-κB),protein kinase B (AKT),p-AKT,mTOR were detected by Western blotting. Results: The degree of stenosis of normal control group was (14.02±2.86)%, saline group was (64.14±3.21)%, penicillin group was (49.11±2.96)%, budesonide group was (39.52±2.09)%, erythromycin group was (32.60±4.27)%. The differences between any two groups were statistically significant (all P<0.05). Except between erythromycin group and normal control group, the differences in relative expression of IL-6 mRNA between any two groups (1.00±0.00, 9.02±1.50, 4.25±0.87, 2.53±0.17, 1.31±0.56) was statistically significant (all P<0.05), and the differences in relative expression of TGF-ß mRNA among all groups (1.00±0.00, 6.92±0.84, 3.83±0.44, 2.13±0.25, 1.40±0.15) were statistically significant (all P<0.05). The relative expression of SIRT-1 mRNA among all the groups (1.000±0.000, 0.209±0.042, 0.375±0.034, 0.555±0.028, 0.667±0.032) was statistically significant different (all P<0.05); except between erythromycin group and budesonide group,the protein levels of SIRT-1 among all other groups (16.93±2.28, 4.77±1.45, 7.70±0.61, 10.76±1.04, 11.03±1.10) were statistically significant different (all P<0.05). The protein levels of mTOR (9.28±4.56, 58.18±8.12, 44.75±5.56, 32.82±5.99, 24.73±3.56) and p-AKT (16.57±4.86, 61.79±6.66, 42.98±5.99, 32.79±5.34, 24.00±4.40) determined through immunohistochemistry of all groups were statistically significant different (all P<0.05). The protein levels of NF-κB, p-NF-κB, AKT, p-AKT and mTOR determined through Western blotting had the same trend as that of determined through immunohistochemistry. The protein expression of NF-κB,AKT and mTOR in saline group were significantly higher than other groups; those protein expression of erythromycin group was lower than budesonide group and penicillin group. Except between the erythromycin group and the normal control group, the protein expression of mTOR in other groups was statistically significant different (all P<0.05). Conclusion: Penicillin,erythromycin and budesonide can alleviate inflammation by increasing SIRT-1, alleviate tracheal scar hyperplasia induced by TGF-beta/mTOR pathway, and reduce the degree of tracheal stenosis in rabbits.

MiR-646 inhibited cell proliferation and EMT-induced metastasis by targeting FOXK1 in gastric cancer.

BACKGROUND: MiR-646 has been reported to be aberrantly expressed in human cancers. However, the underlying molecular mechanisms of action of miR-646 in gastric cancer (GC) have not yet been investigated. METHODS: In vitro function of miR-646 in GC was evaluated using EdU assay, plate colony formation assay, and matrigel invasion assay. Real-time PCR or western blotting was performed to detect miR-646 and FOXK1 expressions. In vivo tumour growth and metastasis were conducted in nude mice. RESULTS: MiR-646 expression was downregulated in GC tissues compared with adjacent normal tissues. Low miR-646 expression is associated with malignant progression. Transient transfection of GC cells with miR-646 inhibited their growth and migration. Moreover, miR-646 influenced the expression of epithelial-mesenchymal transition (EMT)-associated proteins. TGF-ß1 treatment significantly suppressed the expression of miR-646 and overexpression of this microRNA counteracted the influence of the TGF-ß1-induced EMT phenotype. In terms of the underlying mechanism, miR-646 directly targeted FOXK1. In vivo, it inhibited the FOXK1-mediated proliferation and EMT-induced metastasis. Consistently, inverse correlations were also observed between the expression of miR-646 and FOXK1 in human GC tissue samples. Furthermore, miR-646 regulated Akt/mTOR signalling after FOXK1. CONCLUSIONS: miR-646 inhibited GC cell proliferation and the EMT progression in GC cells by targeting FOXK1.

[Effect of low dose erythromycin on the proliferation of granulation tissue after tracheal injury].

Objective: To investigate the effect of low dose erythromycin on the proliferation of granulation tissue after tracheal injury. Methods: Forty-two rabbits were randomly divided into 7 groups (n=6 each), group A (saline control group), group B (penicillin group), group C (low dose erythromycin group), group D (low dose erythromycin and penicillin group), group E (budesonide group), group F (low dose erythromycin and budesonide group), group G (low dose erythromycin, penicillin and budesonide group). All rabbits received tracheotomy, and the tracheal mucosa was scraped with a nylon brush 20 times for tracheal stenosis model. Rabbits were treated with corresponding drugs from a week before operation to 9 days after operation. The serum concentrations of transforming growth factor - beta 1 (TGF-ß(1)), vascular endothelial growth factor (VEGF), interleukin (IL) -6, IL-8 were determined and the tracheal specimens were harvested for measuring degree of stenosis on the 10th day after operation. Results: Serum concentrations of TGF-ß(1) in group A, B, C, D, E, F and G were (17.6±1.3), (18.2±3.1), (13.0±1.1), (14.0±1.0), (21.0±6.1), (13.6± 3.5), (8.2±1.3) ng/L; VEGF were (88.1±4.1), (85.8±4.3), (58.1±6.3), (56.5±2.4), (87.8±2.8), (57.0±3.7), (34.3±6.7) ng/L; IL-6 were (67.8±4.0), (66.1±3.5), (54.1±4.8), (52.1±3.2), (64.6±4.9), (49.4±4.2), (35.9±3.7) ng/L; IL-8 were (112.8±5.2), (116.6±4.1), (88.0±6.2), (85.5±3.5), (114.4±4.6), (82.6±3.8), (55.9±6.0) ng/L, respectively. The serum concentrations of TGF-ß(1), VEGF, IL-6 and IL-8 in group C, D, F and G were significantly lower than those in group A, B and E (all P<0.05). Compared with the other groups, the serum concentrations in group G were the lowest (all P<0.05). All 42 rabbits had tracheal stenosis with different degrees of proliferation of granulation tissue. The degree of tracheal stenosis in Group A, B, C, D, E, F and G were (53.3±4.4)%, (48.2±5.0)%, (24.3±4.4)%, (29.5±3.2)%, (47.8±6.5)%, (27.9±3.1)%, (15.6±2.0)%, respectively. The degree of tracheal stenosis in group C, D, F and G was significantly lower than that in group A, B and E, which had statistical differences (all P<0.05). Compared with the other groups, the degree of tracheal stenosis in group G was the lowest (all P<0.05). Conclusions: Low dose of erythromycin can effectively inhibit the proliferation of granulation tissue after tracheal injury in rabbits. And it has better effectiveness when combined with other antibiotics and hormone.

[Expression and significance of autophagy in rabbit model of tracheal stenosis].

Objective: To investigate the expression and significance of autophagy in rabbit model of tracheal stenosis. Methods: A total of 18 rabbits were equally divided into 3 groups (blank control group, saline group, erythromycin group) in accordance with the random number table. After rabbit model of tracheal stenosis was established, no treatment was done with blank control group. Saline group was atomized with saline (0.54 mg/kg, 2 times/day), and erythromycin group was fed on erythromycin (7.5 mg/kg, 2 times/day) for 7 days before and 10 days after the operation. On the eleventh day, rabbits were executed, and their trachea were collected. The proportion of collagen fiber area of tracheal lamina propria (LP) and epithelium (EP) was assessed by Masson staining. The mRNA of autophagy associated gene-3 (ATG3) and autophagy associated gene-5 (ATG5) of tracheal mucosa were assessed by Real-Time Polymerase Chain Reaction (RT-PCR). The protein of microtubule-associated protein 1 light chain ß(3) (LC3B), ATG3 and ATG5 were assessed by Western blot. Results: The proportion of collagen fiber area of tracheal LP and EP of blank control group was (6.79±0.67)%, saline group was (40.55±5.40)%, erythromycin group was (27.48±0.43)%. The differences between any two groups was all statistically significant (all P<0.01). The relative value of ATG3 mRNA and ATG5 mRNA in saline group were significantly lower than blank control group (all P<0.01). Those value in erythromycin group were significantly higher than the saline group (all P<0.01). The protein levels of LC3B-Ⅱ/Ⅰ, ATG3 and ATG5 in saline group were significantly lower than blank control group (all P<0.01). After low dose of erythromycin intervention, all the protein levels were significantly higher than the saline group (all P<0.01). Conclusions: The expression of autophagy is decreased in rabbit model of trachea stenosis. Low dose of erythromycin could increase the expression of autophagy and at the same time alleviate the degree of fibrosis of the tracheal mucosa. Autophagy may alleviate tracheal fibrosis through up-regulating its expression level and play a protective role.

[Relationship between expression of HDAC2, IL-8, TNF-α in lung adenocarcinoma tissues and smoking].

OBJECTIVE: To investigate the relationship between the expression of histone acetylation enzyme 2 (HDAC2), interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α) in lung adenocarcinoma tissues and smoking. METHODS: A total of 73 cases of lung adenocarcinoma confirmed by pathological examination after surgical removals were collected in the First Affiliated Hospital and Affiliated Tumor Hospital of Guangxi Medical University from April 2014 to March 2015. All patients received preoperative lung function test. Lung adenocarcinoma and para-cancer tissues were cut by the sharp blade and stored in liquid nitrogen and the sampling time was less than 30 minutes. Smokers were defined as people who had smoked more than 100 cigarettes or inhaled the smoke of cigarettes at least one day a week (more than 15 minutes every day) more than three years. According to the lung function and whether smoking or not, the cases of lung adenocarcinoma were divided into three groups: smoking without chronic obstructive pulmonary disease (COPD) group (33 cases), without smoking and COPD group (19 cases), smoking with COPD group (21 cases). The levels of HDAC2, IL-8 and TNF-α mRNA in lung adenocarcinoma and para-cancer tissues of groups were detected by real-time polymerase chain reaction (PCR) and the expression of HDAC2 protein was detected by Western blotting, and statistical analysis was carried out. RESULTS: The expression of HDAC2, IL-8 and TNF-α in lung adenocarcinoma tissues and TNM stage of lung adenocarcinoma showed no significant differences with respect to age and gender (P>0.05). Compared with the para-cancer tissues of 73 cases, the expression of HDAC2 at mRNA and protein levels in lung adenocarcinoma tissues were significantly lower (t=4.15, 8.006, all P<0.01). and the content of IL-8 and TNF-α at mRNA levels were increased (t=-4.252, -5. 576, all P<0.01). The expression of HDAC2 mRNA and protein in lung adenocarcinoma tissues in smoking without COPD group and smoking with COPD group were significantly lower than in without smoking and COPD group (0.38±0.11, 0.35±0.12 vs 0.45±0.10 and 0.26±0.09, 0.24±0.06 vs 0.33±0.10; all P<0.05), and it was the lowest expression in smoking with COPD group. IL-8 and TNF-α at mRNA levels in lung adenocarcinoma tissues in smoking without COPD group and smoking with COPD group were significantly higher than in without smoking and COPD group (0.96±0.19, 1.10±0.18 vs 0.71±0.13 and 0.62±0.21, 0.64±0.20 vs 0.45±0.14; all P<0.05), and the up-regulation was more obvious in smoking with COPD group. The TNM stage of lung adenocarcinoma in smoking group (smoking without COPD group and smoking with COPD group) was higher than without smoking group (without smoking and COPD group)(P=0.038). CONCLUSION: HDAC2 is down-regulated and IL-8, TNF-α are up-regulated in lung adenocarcinoma tissues. They are influenced by smoking and especially when combined with chronic obstructive pulmonary disease.

Two case reports of Castleman disease with pulmonary involvement.

Castleman disease (CD) is a rare reactive lymphoproliferative disorder, first identified in 1954. We recently had the opportunity to analyse the characteristics of two variations of CD with pulmonary involvement. Case 1 had localised retroperitoneal hyaline vascular type CD, while Case 2 was diagnosed as multicentric plasma cell type CD. Both patients had pulmonary symptoms and signs, including cough, dyspnoea, hypoxaemia and ventilatory dysfunction; however, they had different physiological manifestations of their pulmonary abnormalities.

Effects of dietary gossypol concentration on growth performance, blood profiles, and hepatic histopathology in meat ducks.

The objective of this study was to determine the effects of gossypol from cottonseed meal (CSM) on growth performance, blood biochemical profiles, and liver histopathology of ducks. A total of 900 1-d-old ducks were randomly allocated to 5 treatments with 12 pens/treatment and 15 ducks/pen. The 5 experimental diets were formulated in such a way that 0% (a corn-soybean meal basal diet, diet 1), 25% (diet 2), 50% (diet 3), 75% (diet 4), and 100% (diet 5) of protein from soybean meal were replaced with that from CSM. All diets were formulated on a digestible amino acid basis. The experiment included 2 phases, the starter phase (1 to 3 wk) where the test diets contained graded levels of CSM and the growth phase (4 to 5 wk) where birds were fed a corn-soybean basal diet to examine the recovery of ducks after CSM withdrawal. Dietary CSM and gossypol linearly (P < 0.01) and quadratically (P < 0.01) decreased ADG and ADFI during d 1 to 14. The threshold of daily total gossypol (TG) and free gossypol (FG) intake based on ADG on d 1 to 7 and d 7 to 14 were 32.20 and 2.64 mg/d, and 92.12 and 9.62 mg/d, respectively. Serum alanine aminotransferase increased (P < 0.05) linearly with increasing level of gossypol in the diets (d 7), whereas aspartate aminotransferase increased (P < 0.05) linearly and quadratically (d 14). Serum albumin concentration decreased (P < 0.05) quadratically with increasing dietary CSM concentrations on d 21. The degree of damage to the liver increased markedly with increasing dietary CSM and gossypol content and the length of CSM and gossypol intake. On d 35, there was no difference on BW and blood profiles of ducks among all treatments. These results suggest that meat ducks' dietary TG and FG concentration should be lower than 928.9 and 77.2 mg/kg, respectively, during d 1 to 21 of age and that a 2-wk withdrawal of diets containing gossypol should be considered.

A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-â1

We have demonstrated that a synthetic DNA enzyme targeting early growth response factor-1 (Egr-1) can inhibit neointimal hyperplasia following vascular injury. However, the detailed mechanism of this inhibition is not known. Thus, the objective of the present study was to further investigate potential inhibitory mechanisms. Catalytic DNA (ED5) and scrambled control DNA enzyme (ED5SCR) were synthesized and transfected into primary cultures of rat vascular smooth muscle cells (VSMCs). VSMC proliferation and DNA synthesis were analyzed by the MTT method and BrdU staining, respectively. Egr-1, TGF-â1, p53, p21, Bax, and cyclin D1 expression was detected by RT-PCR and Western blot. Apoptosis and cell cycle assays were performed by FACS. Green fluorescence could be seen localized in the cytoplasm of 70.6 ± 1.52 and 72 ± 2.73 percent VSMCs 24 h after transfection of FITC-labeled ED5 and ED5SCR, respectively. We found that transfection with ED5 significantly inhibited cultured VSMC proliferation in vitro after 24, 48, and 72 h of serum stimulation, and also effectively decreased the uptake of BrdU by VSMC. ED5 specifically reduced serum-induced Egr-1 expression in VSMCs, further down-regulated the expression of cyclin D1 and TGF-â1, and arrested the cells at G0/G1, inhibiting entry into the S phase. FACS analysis indicated that there was no significant difference in the rate of apoptosis between ED5- and ED5SCR-transfected cells. Thus, ED5 can specifically inhibit Egr-1 expression, and probably inhibits VSMC proliferation by down-regulating the expressions of cyclin D1 and TGF-â1. However, ED5 has no effect on VSMC apoptosis.

A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-beta1.

We have demonstrated that a synthetic DNA enzyme targeting early growth response factor-1 (Egr-1) can inhibit neointimal hyperplasia following vascular injury. However, the detailed mechanism of this inhibition is not known. Thus, the objective of the present study was to further investigate potential inhibitory mechanisms. Catalytic DNA (ED5) and scrambled control DNA enzyme (ED5SCR) were synthesized and transfected into primary cultures of rat vascular smooth muscle cells (VSMCs). VSMC proliferation and DNA synthesis were analyzed by the MTT method and BrdU staining, respectively. Egr-1, TGF-beta1, p53, p21, Bax, and cyclin D1 expression was detected by RT-PCR and Western blot. Apoptosis and cell cycle assays were performed by FACS. Green fluorescence could be seen localized in the cytoplasm of 70.6 +/- 1.52 and 72 +/- 2.73% VSMCs 24 h after transfection of FITC-labeled ED5 and ED5SCR, respectively. We found that transfection with ED5 significantly inhibited cultured VSMC proliferation in vitro after 24, 48, and 72 h of serum stimulation, and also effectively decreased the uptake of BrdU by VSMC. ED5 specifically reduced serum-induced Egr-1 expression in VSMCs, further down-regulated the expression of cyclin D1 and TGF-beta1, and arrested the cells at G0/G1, inhibiting entry into the S phase. FACS analysis indicated that there was no significant difference in the rate of apoptosis between ED5- and ED5SCR-transfected cells. Thus, ED5 can specifically inhibit Egr-1 expression, and probably inhibits VSMC proliferation by down-regulating the expressions of cyclin D1 and TGF-beta1. However, ED5 has no effect on VSMC apoptosis.

Epithelial neutrophil-activating peptide-78 recruits neutrophils into pleural effusion.

The aim of this study was to investigate the presence of epithelial neutrophil-activating peptide (ENA)-78 in pleural effusions, as well as the chemoattractant activity of pleural ENA-78 on neutrophils. Pleural effusion and serum samples were collected from 75 patients who presented to the respiratory institute (19 with malignant pleural effusion, 21 with tuberculous pleural effusion, 18 with infectious pleural effusion and 17 with transudative pleural effusion). The concentrations of ENA-78, myeloperoxidase and neutrophil elastase were determined, and the chemoattractant activity of ENA-78 for neutrophils both in vitro and in vivo was also observed. The concentrations of ENA-78, myeloperoxidase and neutrophil elastase in infectious pleural effusion were significantly higher than those in malignant, tuberculous and transudative groups, respectively (all p<0.01). Infectious pleural fluid was chemotactic for neutrophils in vitro and anti-ENA-78 antibody could partly inhibit these chemotactic effects. Intrapleural administration of ENA-78 produced a marked progressive influx of neutrophils into pleural space. Compared with noninfectious pleural effusion, ENA-78 appeared to be increased in infectious pleural effusion. Our data suggested that ENA-78 was able to induce neutrophil infiltration into pleural space and might be responsible for pleural neutrophil degranulation.

Effect of inhaled interleukin-4 on airway hyperreactivity in asthmatics.

lnterleukin-4 (IL-4) has been shown to play a crucial role in the pathogenesis of allergic disease including bronchial asthma. In order to investigate the role of IL-4 in airway hyperreactivity, we investigated the effect of inhaled recombinant human IL-4 on airway responsiveness to methacholine and eosinophil numbers in induced sputum in eight patients with allergic asthma using a placebo-controlled study design. Our results demonstrated that in the control experiments receiving vehicle inhalation, methacholine PC20 values did not change nor did the numbers of eosinophils in sputum change from baseline values. In contrast, after IL-4 inhalation, methacholine PC20 fell from baseline (0.43 +/- 1.81 mg/mI) to 0.22 +/- 1.73 mg/mI (p < 0.01) at 24 h, and to 0.21 +/- 1. 74 mg/ml (p < 0.01) at 48 h. Accompanying this increased airway sensitivity was a significant eosinophilia in sputum. Our data indicated that IL-4 increases airway responsiveness by recruiting eosinophils into the airway in patients with allergic bronchial asthma.

Computed tomography in predicting the efficacy of oral cholelitholysis with bile acids.

The efficacy of oral cholelitholytic therapy with chenodeoxycholic acid (CDCA) and ursodeoxycholic acid (UDCA) in 137 patients with gallstones was studied in relation to their CT patterns. The best dissolving results were obtained in patients with the stones in isodensity and faint category (< 50 Hu) on CT. The stones with high density or heterogeneous calcification on CT were insoluble, and therefore, were contraindicated for oral cholelitholytic therapy. The attenuation value of stones was classified as complete dissolution ranged from -2 to 35 Hu (14 +/- 12 Hu, n = 13), and their upper limit of 95 percentiles was 33.4 Hu. CT analysis improved the predictability of dissolving gallstones in comparison with plain abdominal radiography or oral cholecystography (OCG). The complete dissolving rate increased from 9.49% (13/137) in patients selected by classic X-ray to 40.7% (11/27) in isodensity category on CT. Besides, radiolucent gallstones, which showed no obvious filling defect on OCG but distinct echo and shadow on B-type ultrasonography, were also insoluble.