BACKGROUND: It was reported that the reason why the fracture healing become more and more difficult with the increasing age may be associated with the varied gene regulation of bone marrow mesenchymal stem cells differentiation into osteoblast and lipoblast. However, the exact mechanism under it remains poorly understood.OBJECTIVE: To explore the association of the age-related changes on expression of peroxisome proliferator-activated receptor γ (PPARγ) and core binding α1 (Cbfα1) with the age-relatad fracture healing impairment, we studied the expression changes of PPARγ and Cbfα1 during aging fracture healing.DESIGN, TIME AND SETTING: A randomized control animal trial was performed in the Third Xiangya Hospital of Central South University between October 2007 and Febuary 2008. MATERIALS: Six male SD rats of 3 months and the other six of 23 months, randomly-selected, were divided into two groups: the old experiment group (23 months) and the young control group (3 months). METHODS: Transverse osteotomies on the middle-upper parts of left tibiae were performed to the rats, with the self-made extemal mini-fixator implanted there simultaneously. Tractions were taken twice a day (0.2 mm/d) for 14 days from day 2 till day 15 post operation. On day 15 post operation, rats were sacrificed to harvest left tibea samples. MAIN OUTCOME MEASURES: Both imaging examination and histological observation were performed to the fractures of rats. The PPARγ and Cbfα1 expression were detected with reverse transcription-polymerase chain reaction (RT-PCR) technique. RESULTS: All samples were involved in the results analyses. The imaging examination showed that many osteotylus generated between the broken ends of fractures in the young control group; that the bone formation of the young control group was much better than that of the old experiment group. The histological observation showed that large amounts of osteotylus growth occurred to every animal in the young control group and their membrane bone formation was remarkable; comparatively, the osteotylus growth in the old group was weakened dramatically, and only fibrous joints were seen. The RT-PCR detection showed that differences of significance existed in both PPARγ and Cbfα1 mRNA expressions between the two groups, with the much higher Cbfol mRNA expression in the bone marrow of rats in the young group and the much higher PPARy mRNA expression in the bone marrow of rats in the old group.CONCLUSION: Rats of different ages show different fracture healing abilities, i.e. the higher ages, the lower fracture healing abilities. In addition, expressions of PPARγ and Cbfα1 in the bone marrow of rats alter with the increasing age, which indicates that there are certain associations between the changed expression level of the two factors and the fracture healing impartment.

Objective To prepare poly (lactic-co-glycolic acid) (PLGA) microspheres that contain recombinant epidermal growth factor and then evaluate the effect of these sustained release in vitro.Methods Human serum albumin (HAS) microspheres were prepared with a double-emulsification solvent evaporation method.Experiment was designed to optimize the preparation condition of recombinant human epidermal growth factor (rhEGF)-loaded microspheres.Characters of optimal rhEGF-loaded microspheres were analyzed.In-vitro dissolution tests were performed on the microsperes.Results The speed of first emulsification,the concentration of PLGA,the concentration of polyvinyl alcohol (PVA),and the ratio of inner water to organic phase affected the particle size and the encapsulation ratio.The polymer's individual specificities especially density,molecular weight,polymerization,the diameter of microsphere,and the encapsulation efficiency were the crucial factors to influence the speed and time of drug release from microspheres in vitro.The optimal microspheres possessed a smooth and round appearance.It also showed good lente liberantes effect in vitro.Conclusions The optimal microspheres possessed a smooth and round stable appearance and showed good lente liberantes effect in vitro.This technique was simple and had a good reproducibility.

It was found that psoralen derivative could perform a Friedel-Crafts acylation smoothly with acetic anhydride to give 5'-acetylpsoralen in a 73% yield. In the presence of boron trifluoride etherate, 5'-acetylpsoralen reacted with both aromatic amines and aliphatic amine smoothly to afford 5'-Schiff-base group substituted psoralen derivatives in 72%-92% yields. The novel synthetic method has the advantages of cheap materials, mild reaction conditions, good yields and high regioselectivity in the Friedel-Crafts acylation. Cell viability assay by MTT demonstrates that some of the psoralen derivatives 6 have antiproliferative activities.

Objective To construct in the extracellular matrix metalloproteinase inducer (EMMPRIN)glycosylation single point mutation plasmid,and to explore its relationship with tumor cell proliferation.Methods PCR point mutantion technology was used to construct the mutantion plasimid of EMMPRIN glycosylation single point.After successful mutation, the function of mutantion plasmids were detected. Western blotting was used to detect the expression of EMMPRIN protein,immunofluorescence method was used to detemine the morphological changes of the cells,and MTT assay was performed to detect the relationship between mutantion pasmid and tumor cell proliferation.Results Confirmed by restriction enzyme digestion and sequencing,the 44th,the 152th,and the 186th Asn were successfully mutated to Gln in the sequence of EMMPRIN;EMMPRIN/GFP (N44Q), EMMPRIN/GFP(N152Q),and EMMPRIN/ GFP (N186Q)glycosylation single point mutation plasmids were constructed.Compared with wild-type,thel morphology of the cells was significantly changed,the core division of mutant-type cells was significantly reduced, the number of filopodia was reduced. The results of MTT assay showed that the survival rate of the cells in wild-type group were significantly increased compared with control group (P<0.05 );the survival rates of the cells in EMMPRIN (N44Q) group, EMMPRIN (N152Q) group and EMMPRIN(N186Q)were significantly decreased compared with wild-type group(P<0.05).Conclusion Mutant-type EMMPRIN can inhibit the proliferation of tumor cells;with the duration increasing, the inhibitory effect is weakened.There is a correlation between EMMPRIN glycosylation and proliferation of tumor cells.

PURPOSE: This study aimed to identify potential epidermal growth factor receptor (EGFR) gene mutations in non-small cell lung cancer that went undetected by amplification refractory mutation system-Scorpion real-time PCR (ARMS-PCR). MATERIALS AND METHODS: A total of 200 specimens were obtained from the First Affiliated Hospital of Guangzhou Medical University from August 2014 to August 2015. In total, 100 ARMS-negative and 100 ARMS-positive specimens were evaluated for EGFR gene mutations by Sanger sequencing. The methodology and sensitivity of each method and the outcomes of EGFR-tyrosine kinase inhibitor (TKI) therapy were analyzed. RESULTS: Among the 100 ARMS-PCR-positive samples, 90 were positive by Sanger sequencing, while 10 cases were considered negative, because the mutation abundance was less than 10%. Among the 100 negative cases, three were positive for a rare EGFR mutation by Sanger sequencing. In the curative effect analysis of EGFR-TKIs, the progression-free survival (PFS) analysis based on ARMS and Sanger sequencing results showed no difference. However, the PFS of patients with a high abundance of EGFR mutation was 12.4 months [95% confidence interval (CI), 11.6−12.4 months], which was significantly higher than that of patients with a low abundance of mutations detected by Sanger sequencing (95% CI, 10.7−11.3 months) (p < 0.001). CONCLUSION: The ARMS method demonstrated higher sensitivity than Sanger sequencing, but was prone to missing mutations due to primer design. Sanger sequencing was able to detect rare EGFR mutations and deemed applicable for confirming EGFR status. A clinical trial evaluating the efficacy of EGFR-TKIs in patients with rare EGFR mutations is needed.
Adenocarcinoma/genetics ; Adenocarcinoma/pathology ; Aged ; Aged, 80 and over ; Animals ; Base Sequence ; Disease-Free Survival ; Female ; Humans ; Lung Neoplasms/genetics ; Lung Neoplasms/pathology ; Male ; Middle Aged ; Mutation/genetics ; Mutation Rate ; Real-Time Polymerase Chain Reaction/methods ; Receptor, Epidermal Growth Factor/genetics ; Sequence Analysis, DNA/methods ; Treatment Outcome

Objective To describe and introduce the initial clinical application of a novel instrument needle-perc for percutaneous nephrolithotomy (PCNL) in upper urinary tract stones.Methods 24 patients with upper urinary stone treated by PCNL were collected retrospectively between August 2017 and January 2018.Sixteen patients were male and 8 were female.Average age was 41.2 years,ranging 26-65 years.Eight cases had upper pole stones,6 cases had pelvic stones,8 cases had lower pole stones and 4 cases had the stone in UPJ.The mean calculus size was 1.2 cm,ranging 0.5-1.4 cm.All patients were punctured under total ultrasound with needle-perc.Six cases had upper calyceal puncture,10 cases had middle calyceal puncture and 8 cases had lower calyceal puncture.The needle-shaped nephroscope consists of a puncture sheath and a needle handle.The puncture sheath is a hollow metal sheath with an outer diameter of F4.2,an inner diameter of F3.6,and a length of 15 cm.The tip of the sheath is beveled to facilitate puncture.The outer end of sheath is connected to the needle handle through a screw interface.And the three interfaces of the three-way tube can be respectively connected with a liquid irrigation device,a video optical fiber and a 200 μm holmium laser fiber.The needle-perc integrated image system,the irrigation system,and the nephroscope channel are integrated.The tissue passing through the needle can be simultaneously observed through video optical fiber during puncturing.After the tip of the sheath is inserted into the target calyx,the holmium laser fiber is connected for fragmenting or dusting.Results Needle-perc was successful in 22 cases,2 patients were converted to larger tract(F16).The mean opeartive time was 49.2 min,ranging 22-75 min and the mean hemoglobin loss was 5.2 g/L,ranging 0-13.8 g/L.Mean postoperative hospital stay was 3 days,ranging 1 to 6 days.No Double-J stents or nephrostomy tube was placed in the 22 patients.Complications (Clavien Ⅱ) occurred in 4 cases,including fever in 2 cases and renal colic in 2 cases.Plain film of KUB or CT scan was done and stone free rate at 1 month was 90.9％ (20/22),2 patients needed ESWL to remove the residual stones.Conclusions Needle-perc is efficient and safe for small renal stones (size ＜ 1.5 cm) from our initial experience,with high stone-free rate and low complication rate in early follow-up.

Objective To study the effect of diffusion-weighted magnetic resonance imaging (DW-MRI) in the differential diagnosis of hydronephrosis and pyonephrosis.Methods From March 2015 to October 2017,50 patients with renal stone and highly suspected infectious hydronephrosis underwent renal DW-MRI,and clinical materials were collected and analyzed retrospectively.Seventeen male and 33 female patients were enrolled with a mean age of (49.40 ±-10.51) years.The median maximum diameter of renal stone was 30.30 (17.38,56.01) mm and hydronephrosis was 46.39 (34.33,56.55) mm.No pyonephrosis was diagnosed by preoperative ultrasound or CT,while 29 cases of hydronephrosis and 21 cases of pyonephrosis were reported by preoperative DW-MRI.Final diagnoses of hydronephrosis and pyonephrosis were made according to whether the pelvic urine drainage was purulent or not during the surgery.DW-MRI reports and characteristics of DW images were analyzed retrospectively.Apparent diffusion coefficient (ADC) was calculated and ADC map constructed,which was compared between the two groups.Receiver operating characteristic curve (ROC) was drawn to analyze the area under curve (AUC) and the optimal cutoff of ADC value,with sensitivity and specificity.Results Thirty-three patients of hydronephrosis and 17 pyonephrosis were confirmed intraoperatively.The overall accuracy of diagnosis using DW-MRI was 84.00％ (42/50),with sensitivity of 88.24％ (15/17) and specificity of 81.82％ (27/33).Among 30 patients who underwent CT scan in our hospital,mean CT value of 18 hydronephrosis was (7.03 ± 3.26)HU and that of 12 pyonephrosis was (8.67 ± 3.52) HU,with no statistical significance (P ＞ 0.05).On DW image,when b ≥ 500 s/mm2,hydronephrosis signal intensity was lowered apparently,whereas pyonephrosis signal intensity was intensified.On ADC map,hydronephrosis appeared as hyperintensity,whereas pyonephrosis appeared as hypointensity.The mean ADC value of pyonephrosis group was lower than hydronephrosis group [(1.53 ±0.58) × 10-3 mm2/s vs.(2.86 ±0.56) × 10-3 mm2/s,p ＜0.01].ROC analysis revealed that AUC =0.92 (95％ CI 0.80-0.98),and the cut-off value of ADC for pyonephrosis diagnosis was 1.39 × 10-3 mm2/s,of which the sensitivity and specificity was 94.12％ and 84.85％ respectively.Conclusions Signal of pyonephrosis was enhanced on DW image,while decreased on ADC image.The ADC value of pyonephrosis was much lower than that of hydronephrosis,with which the differential diagnosis between pyonephrosis and hydronephrosis could be made efficiently.

Purpose/Significance To explore the characteristics and clinical significance of differentially expressed genes closely re-lated to HPV E6/E7 by using bioinformatics.Method/Process The cervical tissue and clinical information of cervical cancer in TCGA and GTEx of UCSC are used as the training set.The expression profile chip GSE63514 related to cervical cancer in GEO is used as the validation set.Firstly,the limma package of R software is used to screen DEGs of tumor and normal samples,and Venn map of genes re-lated to E6/E7 protein in MigDB is made.Survival analysis is performed by survival kit and verified by ROC and protein expression lev-els.Secondly,key genes are obtained by copy number variation and methylation correlation.Finally,the specific co-expression network is constructed and enrichment analysis and immune infiltration analysis are performed.Result/Conclusion There are 101 differentially expressed genes related to HPV E6/E7 protein,and three genes are found to have significance after screening,namely E2F1,MCM4 and PCNA.At the same time,it is found that the genes in the specific coexpression network are significantly enriched in the DNA replication and chromosome organization pathways.Immune correlation analysis shows that key genes are significantly associated with CD4 T cells,B cells and neutrophils.DNA replication,chromosome organization,etc.,are the molecular mechanisms and key genes significantly related to the development of cervical squamous cell carcinoma and HPV E6/E7 encoded proteins.

Objective:To discuss the regulatory effect of berberine(BBR)on fatty acids in the human glioma T98G cells and its effect on the cell proliferation,migration,and invasion,and to clarify its potential mechanism.Methods:The T98G cells at logarithmic growth phase were divided into control group and different concentrations(25,50,and 100 mg·L-1)of BBR groups.Cell wound healing assay was used to detect the migration rates of the cells in various groups;Transwell chamber assay was used to detect the invasion rates of the cells in various groups.The T98G cells at logarithmic growth phase were divided into control group and 100 mg·L-1 BBR group,and Mass spectrometry was used to detect the fatty acid contents in the cells in two groups.The T98G cells at logarithmic growth phase were divided into control group and different concentrations(50,100,and 150 mg·L-1)of BBR groups;Western blotting method was used to detect the expression levels of phosphatidylinositol 3-kinase(PI3K),phosphorylated PI3K(p-PI3K),protein kinase B(AKT),phosphorylated AKT(p-AKT),sterol regulatory element-binding protein 1(SREBP-1),and fatty acid synthase(FASN)in the cells in various groups.The expression of FASN was suppressed by gene silencing technology,and the T98G cells at logarithmic growth phase were divided into control group,shFASN1 group,and shFASN2 group.Western blotting method was used to detect the expression levels of FASN protein in the cells in various groups;clone formation assay was used to detect the clone formation of the cells in various groups;cell wound healing assay was used to detect the migration rates of the cells in various groups.Results:Compared with control group,the migration rates and invasion rates of the cells in different concentrations of BBR groups were decreased in a concentration-dependent manner(P<0.01),and the fatty acid content in the cells in 100 mg·L-1 BBR group was significantly decreased(P<0.01).Compared with control group,the expression levels of p-PI3K,p-AKT,SREBP-1,and FASN proteins in the cells in 150 mg·L-1 BBR group were significantly decreased(P<0.05 or P<0.01),and the expression level of SREBP-1 protein in the cells in 100 and 150 mg·L-1 BBR groups were significantly decreased(P<0.01).After suppression of FASN expression,compared with control group,the expression levels of FASN protein in the cells in shFASN1 and shFASN2 groups were significantly decreased(P<0.01),and the expression level of FASN protein in the cells in shFASN2 group was lower than that in shFASN1 group(P<0.05);compared with control group,the numbers of clone formation and migration rates of the cells in shFASN1 and shFASN2 groups were significantly decreased(P<0.01),and the migration rate of the cells in shFASN2 group was significantly lower than that in shFASN1 group(P<0.05).Conclusion:BBR interferes with fatty acid synthesis in the glioma T98G cells by reducing the expression of the PI3K/AKT/SREBP-1/FASN pathway related proteins,and decrease their migration and invasion capabilities.

Objective:To quantitatively analyze the plaque components of carotid artery through energy spectrum computed tomography angiography(CTA),and to measure the blood flow perfusion in the blood-supply area of carotid artery through CT perfusion(CTP),so as to explore the relationship among plaque component,the degree of luminal stenosis and cerebral blood flow perfusion.Methods:A total of 68 patients with unilateral plaques of carotid artery and severe vascular stenosis who were screened and diagnosed by ultrasound and CTA in Xiyuan Hospital from December 2017 to July 2019 were selected,and all patients underwent CTA examination and CTP examination.North American symptomatic carotid endarterectomy test(NASCET)method was used to measure the degree of carotid stenosis.The GE AW 4.7 post-process workstation was used to conduct analyses of energy spectrum and cerebral perfusion for the plaque component.And then,the slope of energy spectrum curve and the effective atomic number were obtained.At the same time,the cerebral blood volume(CBV),cerebral blood flow(CBF),time to peak(TTP)and mean transit time(MTT)of contrast agent in blood-supplying area of anterior cerebral artery(ACA)and middle cerebral artery(MCA)at the side of lesion were measured.Results:A total of 68 measured plaques of 68 patients met the condition,including 44 vulnerable plaques(including lipid plaques and mixed plaques)and 24 stable plaques(fibrous plaques).The average slopes of the energy spectrum curves of vulnerable plaque and stable plaque were respectively 0.45±0.45 and 1.15±0.39,and the differences were significant(t=2.413,P<0.05).The averagely effective atomic numbers of vulnerable plaques and stable plaques were respectively 7.21±1.06 and 8.01±0.63,and the difference were significant(t=2.548,P<0.05).The average TTP values of the ACA at the side of lesion of vulnerable plaques and stable plaques were respectively(12.20±1.61)S and(13.59±2.79)S,and the difference was significant(t=-2.607,P<0.05).The mean MTT values of the ACA at the side of lesion of vulnerable plaques and stable plaques were respectively(5.07±1.66)S and(6.09±2.19)S,and the difference was significant(t=-2.177,P<0.05).The degree of vascular stenosis at the side of lesion was positively correlated with TTP and MTT in blood-supplying area of middle cerebral artery(MCA)at the side of lesion(r=0.537,0.465,P<0.05),and that was negatively correlated with CBF values in blood-supplying areas of ACA and MCA at the side of lesion(r=-0.281,-0.569,P<0.05),respectively.The slope of the energy spectrum curve of carotid plaque was positively correlated with the TTP values in blood-supplying areas of ACA and MCA at the side of lesion(r=0.242,0.246,P<0.05),respectively.Conclusion:CT spectral imaging can quantitatively analyze the displayed components of carotid atherosclerotic plaque,and the degree of vascular stenosis can affect the blood flow perfusion of cerebral tissue,and the delays of TTP and MTT are more easily caused by vulnerable plaque,and the TTP of them is more sensitivity.