[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.

[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.

Disease correction by combined neonatal intracranial AAV and systemic lentiviral gene therapy in Sanfilippo Syndrome type B mice.

Mucopolysaccharidosis type IIIB (MPS IIIB) or Sanfilippo Syndrome type B is a lysosomal storage disease resulting from the deficiency of N-acetyl glucosaminidase (NAGLU) activity. We previously showed that intracranial adeno-associated virus (AAV)-based gene therapy results in partial improvements of several aspects of the disease. In an attempt to further correct the disease, MPS IIIB mice were treated at 2-4 days of age with intracranial AAV2/5-NAGLU (IC-AAV), intravenous lentiviral-NAGLU (IV-LENTI) or the combination of both (BOTH). The BOTH group had the most complete biochemical and histological improvements of any treatment group. Compared with untreated MPS IIIB animals, all treatments resulted in significant improvements in motor function (rotarod) and hearing (auditory-evoked brainstem response). In addition, each treatment group had a significantly increased median life span compared with the untreated group (322 days). The combination arm had the greatest increase (612 days), followed by IC-AAV (463 days) and IV-LENTI (358 days). Finally, the BOTH group had nearly normal circadian rhythm measures with improvement in time to activity onset. In summary, targeting both the systemic and central nervous system disease of MPS IIIB early in life appears to be the most efficacious approach for this inherited metabolic disorder.

Learning chronobiology by improving Wikipedia.

Although chronobiology is of growing interest to scientists, physicians, and the general public, access to recent discoveries and historical perspectives is limited. Wikipedia is an online, user-written encyclopedia that could enhance public access to current understanding in chronobiology. However, Wikipedia is lacking important information and is not universally trusted. Here, 46 students in a university course edited Wikipedia to enhance public access to important discoveries in chronobiology. Students worked for an average of 9 h each to evaluate the primary literature and available Wikipedia information, nominated sites for editing, and, after voting, edited the 15 Wikipedia pages they determined to be highest priorities. This assignment (http://www.nslc.wustl.edu/courses/Bio4030/wikipedia_project.html) was easy to implement, required relatively short time commitments from the professor and students, and had measurable impacts on Wikipedia and the students. Students created 3 new Wikipedia sites, edited 12 additional sites, and cited 347 peer-reviewed articles. The targeted sites all became top hits in online search engines. Because their writing was and will be read by a worldwide audience, students found the experience rewarding. Students reported significantly increased comfort with reading, critiquing, and summarizing primary literature and benefited from seeing their work edited by other scientists and editors of Wikipedia. We conclude that, in a short project, students can assist in making chronobiology widely accessible and learn from the editorial process.