Long-term hypertension causes excessive vascular remodeling and leads to adverse cardiovascular events. Balance of ubiquitination and deubiquitination has been linked to several chronic conditions, including pathological vascular remodeling. In this study, we discovered that the expression of ubiquitin-specific protease 25 (USP25) is significantly up-regulated in angiotensin II (Ang II)-challenged mouse aorta. Knockout of Usp25 augments Ang II-induced vascular injury such as fibrosis and endothelial to mesenchymal transition (EndMT). Mechanistically, we found that USP25 interacts directly with Forkhead box O3 (FOXO3) and removes the K63-linked ubiquitin chain on the K258 site of FOXO3. We also showed that this USP25-mediated deubiquitination of FOXO3 increases its binding to light chain 3 beta isoform and autophagosomic-lysosomal degradation of FOXO3. In addition, we further validated the biological function of USP25 by overexpressing USP25 in the mouse aorta with AAV9 vectors. Our studies identified FOXO3 as a new substrate of USP25 and showed that USP25 may be a potential therapeutic target for excessive vascular remodeling-associated diseases.

AIM:To investigate the role of lipocalin 2(LCN2)in lipopolysaccharide(LPS)-induced microg-lia polarization in mice and to elucidate the potential mechanisms involving the P38 mitogen-activated protein kinase(MAPK)pathway.METHODS:BV2 microglia were treated with LPS to induce M1 polarization,and short hairpin RNA(shRNA)and exogenous LCN2 protein were used to silence or overexpress LCN2 in BV2 cells.BV2 microglia were cul-tured in vitro and divided into the following groups:control,LPS(100 μg/L),LPS+sh-NC,LPS+sh-LCN2,and LPS+LCN2(1 mg/L).Flow cytometry was used to detect the number of CD16/32+and CD206+T cells.Western blot and RT-qP-CR were employed to measure the protein and mRNA levels of P38 MAPK,PGC-1α,and PPARγ to assess the effects of LCN2 on LPS-induced BV2 cell polarization and the P38 MAPK pathway.Additionally,the P38 MAPK pathway inhibitor SB203580 was used to treat LPS or LCN2-induced cells.The cells were categorized into control,LPS,LPS+LCN2(1 mg/L),LPS+SB203580(50 nmol/L),and LPS+LCN2+SB203580 groups.ELISA was used to measure inflammatory factor levels,Western blot was used to detect M1/M2 marker proteins,and Western blot and RT-qPCR were used to analyze pro-tein and mRNA expressions in the P38 MAPK pathway.RESULTS:LPS significantly increased LCN2 expression in BV2 cells(P＜0.05)and induced M1 polarization(P＜0.01).Silencing LCN2 reduced LCN2 expression and M2 polarization in LPS-induced BV2 cells(P＜0.01),increased M1 polarization(P＜0.01),and inhibited activation of the P38 MAPK-PGC-1α-PPARγ pathway(P＜0.05).Conversely,exogenous addition of LCN2 promoted M2 polarization in LPS-induced BV2 cells and activated the P38 MAPK pathway(P＜0.05).The use of a P38 MAPK pathway inhibitor further confirmed that LCN2 modulates LPS-induced microglia polarization through the P38 MAPK pathway.CONCLUSION:LCN2 inhibits LPS-mediated M1 polarization of BV2 microglia by activating the P38 MAPK pathway,thereby playing a protective role in neuroinflammatory responses.

AIM:To investigate the role of lipocalin 2(LCN2)in lipopolysaccharide(LPS)-induced microg-lia polarization in mice and to elucidate the potential mechanisms involving the P38 mitogen-activated protein kinase(MAPK)pathway.METHODS:BV2 microglia were treated with LPS to induce M1 polarization,and short hairpin RNA(shRNA)and exogenous LCN2 protein were used to silence or overexpress LCN2 in BV2 cells.BV2 microglia were cul-tured in vitro and divided into the following groups:control,LPS(100 μg/L),LPS+sh-NC,LPS+sh-LCN2,and LPS+LCN2(1 mg/L).Flow cytometry was used to detect the number of CD16/32+and CD206+T cells.Western blot and RT-qP-CR were employed to measure the protein and mRNA levels of P38 MAPK,PGC-1α,and PPARγ to assess the effects of LCN2 on LPS-induced BV2 cell polarization and the P38 MAPK pathway.Additionally,the P38 MAPK pathway inhibitor SB203580 was used to treat LPS or LCN2-induced cells.The cells were categorized into control,LPS,LPS+LCN2(1 mg/L),LPS+SB203580(50 nmol/L),and LPS+LCN2+SB203580 groups.ELISA was used to measure inflammatory factor levels,Western blot was used to detect M1/M2 marker proteins,and Western blot and RT-qPCR were used to analyze pro-tein and mRNA expressions in the P38 MAPK pathway.RESULTS:LPS significantly increased LCN2 expression in BV2 cells(P＜0.05)and induced M1 polarization(P＜0.01).Silencing LCN2 reduced LCN2 expression and M2 polarization in LPS-induced BV2 cells(P＜0.01),increased M1 polarization(P＜0.01),and inhibited activation of the P38 MAPK-PGC-1α-PPARγ pathway(P＜0.05).Conversely,exogenous addition of LCN2 promoted M2 polarization in LPS-induced BV2 cells and activated the P38 MAPK pathway(P＜0.05).The use of a P38 MAPK pathway inhibitor further confirmed that LCN2 modulates LPS-induced microglia polarization through the P38 MAPK pathway.CONCLUSION:LCN2 inhibits LPS-mediated M1 polarization of BV2 microglia by activating the P38 MAPK pathway,thereby playing a protective role in neuroinflammatory responses.

Objective To explore the relationship between ileocolonic lesions and perianal fistulas of Crohn's disease as sessed by CT enterography (CTE).Methods Totally 28 patients with initial diagnosis of active ileocolonic lesions of Crohn 's disease were collected,16 with perianal fistula and 11 without perianal fistulas.All patients underwent CTE and pelvic MRI.Total number of lesions,minimum length between every two lesions in colon wall and maximum length of colonic le sions were calculated.The rank sum test was performed respectively.Results Lesions of 14 patients (14/16,87.50％) in perianal fistulas group located in left colon or rectum,while lesions of 6 patients (6/12,50.00 ％) in non-perianal fistulas group located in left colon or rectum,the difference was statistically significant (Z=-2.135,P＜0.05).The mean number of lesions in patients with perianal fistulas was 3.06,while in patients without perianal fistulas was 2.91,there was no statistical difference (P＞0.05).The maximum length of colonic lesions in patients with perianal fistulas was (12.79± 8.30)cm,while in patients without perianal fistulas was (7.04± 3.09)cm,and there was no statistical difference(P＞ 0.05).The minimum length hetween every two lesions in patients with perianal fistulas was (5.23±2.98)cm,while in pa tients without perianal fistulas was (8.44 ± 2.87) cm,the difference was statistically significant (Z =-2.095,P＜ 0.05).Conclusion Crohn's disease complicated with perianal fistulas has relationship with lesion location and smaller length intervals between two lesions in colon wall,and has no relationship with total number of lesions and maximum length of colon lesions.

OBJECTIVETo evaluate the influences of inferior mesenteric artery (IMA) types and Riolan artery arcade absence on the incidence of anastomotic leakage(AL) after laparoscopic resection of rectal cancer.

METHODSClinical data of 116 local advanced rectal cancer patients who underwent laparoscopic resection in The Sixth Affiliated Hospital of Sun Yat-sen University from January 2012 to December 2014 were analyzed retrospectively. IMA and Riolan artery arcade were examined by preoperative computed tomography angiography (CTA) reconstruction. The influences of IMA type, Riolan artery arcade absence and ligation site (high or low) on AL were analyzed by Logistic regression.

RESULTSThe proportion of IMA types(I(-IIII() was 57.8%(67/116), 10.3%(12/116), 31.0%(36/116) and 0.9%(1/116), respectively. Riolan artery arcade absence was found in 60.3%(70/116). Eight (6.9%) patients suffered from AL. IMA type III( had significantly higher AL incidence as compared to other IMA types [19.4%(7/36) vs. 1.2%(1/80), P=0.001]. Meanwhile, patients with Riolan artery arcade absence also had significantly higher AL incidence[11.4%(8/70) vs. 0.0%(0/46), P=0.030]. However, the difference in AL incidence between high and Low IMA ligation was not statistically significant [8.0%(7/87) vs. 3.4%(1/29), P=0.531]. Seven of these 8 AL patients were found in IMA type III( with Riolan artery arcade absence and high ligation. Multivariate analysis showed that IMA type III( (P=0.001) and Riolan artery arcade absence (P=0.002) were independent risk factors of AL.

CONCLUSIONSIMA type III( with Riolan artery arcade absence increases AL incidence significantly in laparoscopic resection of rectal cancer. IMA type and Riolan aretry arcade absence or not contribute to the selection of IMA ligation site in the operation. For the colorectal cancer patients with IMA type III( and Riolan artery arcade absence, selective low IMA ligation with root lymph node dissection should be recommended.

Adult ; Anastomotic Leak ; Arteries ; Colorectal Neoplasms ; surgery ; Female ; Humans ; Incidence ; Laparoscopy ; Ligation ; Lymph Node Excision ; Lymphatic Metastasis ; Male ; Mesenteric Artery, Inferior ; Middle Aged ; Rectal Neoplasms ; surgery ; Retrospective Studies

A two_dimensional HPLC method was developed for the determination of methotrexat ( MTX ) in human plasma. The samples were treated with trichloroacetate for sedimentation and high speed centrifugation, and the obtained supernatant was taken for analysis. The analytes in sample were separated on the first dimension column (ASTON C8 100 mm × 4. 6 mm, 5μm), and trapped on the middle column (ASTON SCX 20 mm × 4. 6 mm, 5 μm) using valve_switching technique for purification and storage. Finally, the trapped analytes were transferred to the second_dimension column (SAC C8 100 mm × 4. 6 mm, 5μm) for the second separation. The mobile phase used for the first dimension was 10 mmol/L ammonium acetate_acetonitrile(9∶1, V/V, pH=3. 8) with a flow rate of 1 mL/min and the mobile phase used for middle column was 10 mmol/L phosphoric acid ( pH=3 . 0 ) . The mobile phase used in second_dimension was a mixed solution of 50 mmol/L ammonium acetate and acetonitrile (87∶10, V/V, pH=5. 2). UV detection was carried out at 306 nm and completed in 4 min. The calibration curve showed a linearity range from 0. 0879 to 5. 154 μmol/L (r=0. 99998). The LOQ was 0. 005 μmol/L. The intra_and inter_day precisions were lower than 1. 5% and 1. 8%, respectively. The relative recovery and the absolute recovery were 99. 1% - 101. 2% and 85. 67%-86. 35%, respectively. The assay is simple, accurate, reproducible, and suitable for the therapeutic drug monitoring of MTX in the hospital and the study on the pharmacokinetics of MTX.

Objective To analyze retrospectively the impact of different heart rates on image quality and radiation dose of coronary angiography using 640-slice dynamic volume CT.Methods A total of 461 patients with suspected coronary artery disease or referred to health check underwent coronary angiography with 640-slice dynamic volume CT.Two groups were created according to their heart rates:Group A had heart rate < 65 beats per minute (n=337)and Group B had heart rate between 65 to 122 beats per minute (n=124).Image quality was assessed by analyzing the 1 5 segments of the main coronary branches using 3-grade scale (grade I-good,grade Ⅱ-acceptable,grade Ⅲ-poor).Effective radiation dose was also evaluated.Results Patients in both A group and B group performed successful CT coronary angiography with a total of 6 91 5 coronary segments.Among them,coronary segments that could be evaluated reliably accounted for 94.5% (6 535/6 91 5)while 5.5% (380/6 91 5)were too small (≤1.5 mm)to be assessed. For the image quality,Group A and Group B had grade I in 90.5% (305/337)vs 74.2% (92 /124),grade Ⅱ in 9.5% (32/337)vs 21.0% (26 / 124)and grade Ⅲ in 0 % (0 / 337 )vs 4.8% (6 / 124),respectively.Image quality was significantly different be-tween Group A and Group B (P <0.001).In addition,32 patients (9.5%)in Group A had slight cardiac motion artifacts but with-out affecting image quality,whereas 26 patients (21.0%)in Group B had higher degree of cardiac motion artifacts thus graded as grade Ⅱ.Stair-step artifacts were not found in all patients.The effective radiation dose was higher in Group B than in Group A by 32.05%(7.91±0.34 mSv vs 5.99±0.17 mSv).Conclusion Coronary angiography using 640-slice dynamic volume CT can guarantee excellent image quality when heart rate < 65 beats per minute.Although the image quality would decrease in some extent it is still diagnostic when heart rate is between 65 to 122(include 65 and 122)beats per minute.

OBJECTIVETo evaluate the application of low-dose CT enterography with adaptive iterative dose reduction(AIDR) technique in diagnosing Crohn's disease.

METHODSRetrospective analysis was performed on 26 patients diagnosed as Crohn's disease by the multidisciplinary team in our hospital. Low-dose CT enterography with 640-slice MDCT was performed on these 26 patients using adaptive iterative dose reduction(AIDR) technique. Characteristics of Crohn's disease in CT enterography images were independently analyzed by two radiologists who were experienced in Crohn's disease with calculating the total radiation dosage.

RESULTSThe radiation dosage of 26 patients ranged from 5.58 to 12.90 [mean (9.00±2.00)] mSv, which was lower than conventional scan (around 15 mSv) known from the literatures. According to the images of CT enterography of 26 cases, bowel wall thickening with abnormal enhancement and lymphadenectasis were found in 25 cases with total 109 segmental bowel wall thickening. Among 25 thickening cases, enterostenosis was found in 16 cases, stratification enhancement in 12 cases and comb sign in 14 cases. Besides, it was found that 8 cases with hyperdense fat on the mesenteric side, 7 cases with intestinal fistula, 6 cases with abdominal cavity abscess, and 3 cases with anal fistula.

CONCLUSIONCT enterography of Crohn's disease with adaptive iterative dose reduction technique is an effective method to evaluate Crohn's disease without compromising image quality with reduced radiation dosage.

Crohn Disease ; diagnostic imaging ; Humans ; Intestinal Fistula ; Radiation Dosage ; Rectal Fistula ; Retrospective Studies ; Tomography, X-Ray Computed ; methods

OBJECTIVE: To investigate whether single nucleotide polymorphisms (SNPs) of rs2298771 and rs3812718 of the sodium channel α-subunit type 1 (SCN1A) gene affect the efficacy of carbamazepine (CBZ) treatment for seizures in Chinese Han epileptic patients. METHODS: SNP rs2298771 and rs3812718 of the SCN1A gene from 628 patients were genotyped. CBZ monotherapy was administered to the subjects with new-onset partial seizures. The efficacy was defined as the decrease in the number of seizures. Four semi-quantitative levels were used to assess the efficacy: seizure-free (SF), >75% seizure decrease (SD), 50%-75% SD, and <50% SD in the number of seizures compared with patients' initial conditions. RESULTS: After the 12 month treatment with CBZ monotherapy, the rate of SF patients with G allele of the SNP rs2298771 was significantly lower than that in patients with the AA genotype (P=0.003). The heterozygote and homozygote of the G allele at SNP rs2298771 predicted the low SF rate (OR=2.101, 95% CI 1.289-3.425). Marginal significance was observed between the dichotomous efficacy of SF and non-SF in 3 partial seizure types (P=0.028). CONCLUSION rs2298771 is significantly associated with the efficacy of CBZ monotherapy in Chinese Han epileptic patients.
Alleles ; Asian Continental Ancestry Group ; Carbamazepine ; therapeutic use ; Epilepsy ; Genotype ; Humans ; NAV1.1 Voltage-Gated Sodium Channel ; genetics ; Polymorphism, Single Nucleotide ; Seizures ; drug therapy ; genetics