Objective To observe the eotaxin expression of rat airway smooth muscle cells ( ASMCs) induced by serum from asthmatic rats, and explore the possible mechanism. Methods ASMCs isolated fromrat tracheas were cultured in vivo. Then they were treated with serum from asthmatic rats, or treated with serum and dexamethasone simultaneously. The level of eotaxin protein in supernatant and eotaxin mRNA in ASMCs were measured by ELISA and reverse transcription-polymerase chain reaction. The expression of cAMP in ASMCs was examined by radioimmunoassay. Results After the treatment with sensitized serum, the eotaxin level in supernatant and mRNA expression in ASMCs were significantly higher [ ( 107. 09 ±7. 12) ng/L vs. ( 0. 63 ±0. 56) ng/L, P lt; 0. 05; 1. 39 ±0. 04 vs. 0. 05 ±0. 01, P lt;0. 05] , and the level of cAMP in ASMCs was significantly lower compared with the control group [ ( 17. 58 ±3. 62) ng/L vs. ( 32. 39 ±3. 36) ng/L, P lt; 0. 05] . After intervened by the sensitized serum and dexamethasone simultaneously, the protein and mRNA expressions of eotaxin were lower compared with those intervened by sensitized serumalone [ ( 64. 18 ±4. 04) ng/L and 0. 77 ±0. 19] . The level of eotaxin in supernatant was negatively correlated with cAMP level in ASMCs ( r = - 0. 788, P lt; 0. 01) . Conclusions There is anautocrine function in ASMCs as inflammatory cells after stimulation with sensitized serum. Eotaxin may play an important roll in the pathogenesis of asthma via a cAMP-dependent pathway.
ObjectiveTo detect the expression of Krüppel like factor 8 (KLF8) in breast cancer tissues and cells and to explore the clinical significance of KLF8.Methods① The Oncomine database was used to analyze the differential expression of KLF8 mRNA in the breast cancer tissues. The Kaplan-Meier Plotter database was used to analyze the relationship between KLF8 mRNA expression and prognosis (relapse free survival, overall survival, post-progression survival, and distant metastasis-free survival) of patients with breast cancer. ② The quantitative real-time PCR (qRT-PCR) and Western blot were used to detect the KLF8 expression levels in the 16 clinical patients with breast cancer and 7 breast cancer cell lines (MDA-MB-231, MCF-12A, Hs-578T, MCF-7, BT-474, MDA-MB-453, ZR-75-30) and normal breast epithelial cell lines MCF-10A, and the immunofluorescence was used to further detect the localization of KLF8 expression in the 2 breast cancer cell lines with higher KLF8 expression level. ③ The immunohistochemistry was used to detect the expression of KLF8 protein in 135 cases of breast cancer tissue microarrays, and the relationships between KLF8 protein expression and clinicopathologic characteristics or overall survival were analyzed.Results① The Oncomine database showed that KLF8 mRNA expression in the breast cancer tissues was higher than that in the normal breast tissues (P<0.001). The median KLF8 mRNA expression level was taken as the cut-off point for high or low KLF8 expression. The results of Kaplan-Meier Plotter data analysis showed that the prognosis (relapse free survival, overall survival, postprogression survival, and distant metastasis-free survival) of patients with low KLF8 mRNA expression were better than those of patients with high KLF8 mRNA expression (P<0.05). ② The results of qRT-PCR and Western blot all showed that the KLF8 mRNA and protein expression levels in the breast cancer tissues were higher than those in the adjacent normal tissues (P=0.002, P<0.001). In addition, the Western blot results showed that the expression of KLF8 protein in the 7 breast cancer cell lines was higher than that in the normal breast epithelial cell lines MCF-10A respectively, and KLF8 protein mainly expressed in the cytoplasm of breast cancer cells and highly expressed in the nuclear of a few cells. ③ There were 63 cases of high KLF8 expression and 72 cases of low KLF8 expression by the immunohistochemical analysis of 135 patients with breast cancer tissue microarray (the H-score of the immunohistochemical test results was 75 as the cut-off point, H-score >75 was the high KLF8 expression and H-score ≤75 was the low KLF8 expression), the differences of statuses of estrogen receptor (ER) and progesterone receptor (PR) between the patient with high KLF8 expression and low KLF8 expression were significant (P<0.05). The Kaplan-Meier survival curve analysis showed that the prognosis of patients with high KLF8 expression was worse than that of patients with low KLF8 expression (P=0.002). The univariate analysis showed that the TNM stage, statuses of ER and PR, and KLF8 expression were related to the prognosis of patients with breast cancer (P<0.05), further multivariate Cox proportional hazards regression analysis indicated that the later stage of TNM and high KLF8 expression were the independent risk factors (P<0.05).ConclusionsThe results of this study suggest that KLF8 highly expresses in both breast cancer tissues and breast cancer cells, which is related to the statuses of ER and PR and prognosis of patients with breast cancer. KLF8 might be involved in the progression of breast cancer as an oncogenic gene, or it might provide a new direction for prognosis judgment and molecular targeted therapy of breast cancer.
The Chinese human hepatocellular carcinoma cell SMMC7721 has been analysed by flow cytometry, Southern blotting and Western blotting. The results indicated that SMMC7721 cell is a hypoploid cell with a 0.81 DNA index, and that SMMC7721 cell has internal deletion in the 5'-end of its Rb gene and has no Rb gene product (Rb protein). The normal Rb cDNA has been inserted into a retrovirus vector DOL and introduced into SMMC7721 cells by electrporation transfection technique.About 75% of transfected SMMC7721 cells have been killed by Rb gene product. The remain 25% cells are alive as exogenous Rb gene has been mutationally inactivated. （Chin J Ocul Fundus Dis,1994,10:21-24）
In recent years, the computer science represented by artificial intelligence and high-throughput sequencing technology represented by omics play a significant role in the medical field. This paper reviews the research progress of the application of artificial intelligence combined with omics data analysis in the diagnosis and treatment of non-small cell lung cancer (NSCLC), aiming to provide ideas for the development of a more effective artificial intelligence algorithm, and improve the diagnosis rate and prognosis of patients with early NSCLC through a non-invasive way.
The biomaterial, chitin, was used to create a nerve regeneration chamber for bridging healing experiment of sciatic nerve of rats having a defect of 12mm. The crude Schwann cells were introduced into the chambers in one group and the other group had no crude Schwann cells in the chamber and the results of the two groups were compared with those having the nerve defects bridged with skeletal muscles. The specimens were observed by macroscopic, microdissection. electrophysiologic testing, HRP retrograde labelling, histologic and electron microscopic examinations at 4, 8, and 12 weeks after the operation. The results showed that atthe 8th week, the regenerating nerve fibers from the cephalad ends had united with the fibers of the caudal ends of the divided nerves either the crude Schwanneclls were introduced or not, but the morphology of the regenerating nerve, the way of regeneration and the recovery of the function of the extremities were far superior in the group that no cruds Schwann cells had been introduced than those with crude Schwann cell introduced and those bridged by skeletal muscles.
Objective To develop a new method for a tissue engineered vascular graft by combining endothelial cells and an acelluarized allogenic matrix. Methods Acellularized matrix tubes were obtained by a 0.1% trypsin and 0 02% EDTA solution for 24 hours and 1% Triton X 100 for 176 hours, respectively. Endothelial cells were isolated from alloaorta and expanded in vitro. Finally, the inner surface of acellularized matrix was reseeded with endothelial cells. Acellularity and reseeding were analysed by light microscopy and scanning electron microscopy. Results The acellularization procedure resulted in an almost complete removal of the original cells and the loose three-dimensional (3D) matrix. The acellular matrix could be reseeded with expanded endothelial cells in vitro, and endothelial cells had the potential of spread and proliferation. Conclusion Acellular matrix produces by Tritoon X-100 and trypsin possesses satisfactory biocompatibility for allogenic endothelial cell. Vascular grafts can be generated in vitro by a combination of endothelial cells and allogenic acelluarized matrix.
OBJECTIVE To review the recent research progress of bone-marrow stromal stem cells (BMSCs) in the conditions of culture in vitro, chondrogenic differentiation, and the application in cartilage tissue engineering. METHODS: Recent original articles related to such aspects of BMSCs were reviewed extensively. RESULTS: BMSCs are easy to be isolated and cultivated. In the process of chondrogenesis of BMSCs, the special factors and interaction between cells are investigated extensively. BMSCs have been identified to form cartilage in vivo. One theory is the committed chondrocyte from BMSCs is only a transient stage. CONCLUSION: BMSCs are the alternative seeding cells for cartilage tissue engineering. The conditions promoting mature chondrocyte should be further investigated.
ObjectiveTo investigate the effect of nicotinamide mononucleotide adenosyl transferase 3 (NMNAT3) on the mitochondrial function and anti-oxidative stress of rabbit bone marrow mesenchymal stem cells (BMSCs) under oxidative stress in vitro by regulating nicotinamide adenine dinucleotide (NAD+) levels.MethodsThe bone marrow of femur and tibia of New Zealand white rabbits were extracted. BMSCs were isolated and cultured in vitro by density gradient centrifugation combined with adherent culture. The third generation cells were identified by flow cytometry and multi-directional induction. Overexpression of NMNAT3 gene was transfected into rabbit BMSCs by enhanced green fluorescent protein (EGFP) labeled lentivirus (BMSCs/Lv-NMNAT3-EGFP), and then the expression of NMNAT3 was detected by real-time fluorescence quantitative PCR (qRT-PCR) and Western blot and cell proliferation by cell counting kit 8 (CCK-8) method. BMSCs transfected with negative lentivirus (BMSCs/Lv-EGFP) and untransfected BMSCs were used as controls. The oxidative stress injury cell model was established by using H2O2 to treat rabbit BMSCs. According to the experimental treatment conditions, they were divided into 4 groups: Group A was normal BMSCs without H2O2 treatment; untransfected BMSCs, BMSCs/Lv-EGFP, and BMSCs/Lv-NMNAT3-EGFP in groups B, C, and D were treated with H2O2 simulated oxidative stress, respectively. The effects of NMNAT3 on the mitochondrial function of BMSCs under oxidative stress [changes of mitochondrial membrane potential, NAD+ and adenosine triphosphate (ATP) levels], the changes of anti-oxidative stress ability of BMSCs [reactive oxygen species (ROS) and malondialdehyde (MDA) levels, manganese superoxide dismutase (Mn-SOD) and catalase (CAT) activities], and the effects of BMSCs on senescence and apoptosis [senescence associated-β-galactosidase (SA-β-gal) staining and TUNEL staining] were detected after 24 hours of treatment.ResultsThe rabbit BMSCs were successfully isolated and cultured in vitro. The stable strain of rabbit BMSCs with high expression of NMNAT3 gene was successfully obtained by lentiviral transfection, and the expressions of NMNAT3 gene and protein significantly increased (P<0.05). There was no significant difference in the trend of cell proliferation compared with normal BMSCs. After treatment with H2O2, the function of mitochondria was damaged and apoptosis increased in all groups. However, compared with groups B and C, the group D showed that the mitochondrial function of BMSCs improved, the membrane potential increased, the level of NAD+ and ATP synthesis of mitochondria increased; the anti-oxidative stress ability of BMSCs enhanced, the levels of ROS and MDA decreased, and the activities of antioxidant enzymes (Mn-SOD, CAT) increased; and the proportion of SA-β-gal positive cells and the rate of apoptosis decreased. The differences in all indicators between group D and groups B and C were significant (P<0.05).ConclusionNMNAT3 can effectively improve the mitochondrial function of rabbit BMSCs via increasing the NAD+ levels, and enhance its anti-oxidative stress and improve the survival of BMSCs under oxidative stress conditions.
ObjectiveTo exclusively compare the short-and long-term outcomes of hepatic resection (HR) patients with multifocal tumors meeting the Milan criteria between locating in same and different sections. MethodsA total of 219 consecutive HR patients with multifocal tumors meeting the Milan criteria were divided into group SS (n=97, same section) and group DS (n=122, different sections) according to their anatomical location (Couinaud's segmentation). ResultsThe 1-, 3-, and 5-year overall survival (OS) and recurrence-free survival (RFS) rates were significantly higher in the group SS than those in the group DS (P < 0.05). The subgroup analysis showed that patients with 2 tumors and those undergoing en bloc resection were associated with better OS and RFS (P < 0.05). ConclusionsFor HCC patients with multifocal tumors meeting the Milan criteria, those with tumors locating in same hepatic section may have better longterm survival and lower HCC recurrence rates than those locating in different sections after HR.
ObjectiveTo investigate the feasibil ity and effectiveness of using scar spl it thickness skin grafts combined with acellular allogeneic dermis in the treatment of large deep Ⅱ degree burn scar. MethodsBetween January 2013 and December 2013, 20 cases of large deep Ⅱ degree burn scar undergoing plastic operation were enrolled. There were 14 males and 6 females, aged 4 to 60 years (mean, 40 years). Burn reasons included hydrothermal burns in 10 cases, flame burns in 9 cases, and lime burns in 1 case. The burn area accounted for 70% to 96% total body surface area (TBSA) with an average of 79% TBSA. The time from wound healing to scar repair was 3 months to 2 years (mean, 7 months). Based on self-control, 0.7 mm scar spl it thickness skin graft was used to repair the wound at the right side of joints after scar resection (control group, n=35), 0.5 mm scar spl it thickness skin graft combined with acellular allogeneic dermis at the left side of joints (trial group, n=30). Difference was not statistically significant in the scar sites between 2 groups (Z=-1.152, P=0.249). After grafting, negative pressure drainage was given for 10 days; plaster was used for immobilization till wound heal ing; and all patients underwent regular rehabil itation exercises. ResultsNo significant difference was found in wound heal ing, infection, and healing time between 2 groups (P>0.05). All patients were followed up for 6 months. According to the Vancouver Scar Scale (VSS), the score was 5.23±1.41 in trial group and was 10.17±2.26 in control group, showing significant difference (t=8.925, P=0.000). Referring to Activities of Daily Living (ADL) grading standards to assess joint function, the results were excellent in 8 cases, good in 20 cases, fair in 1 case, and poor in 1 case in trial group; the results were excellent in 3 cases, good in 5 cases, fair in 22 cases, and poor in 5 cases in control group; and difference was statistically significant (Z=-4.894, P=0.000). ConclusionA combination of scar spl it thickness skin graft and acellular allogeneic dermis in the treatment of large deep Ⅱ degree burn scar is feasible and can become one of solution to the problem of skin source tension.