Objective To explore the diagnostic value of k ras mutation detection of specimen obtained by endoscopic ultrasound-guided fine needle aspiration (EUS-FNA)for pancreatic cancer.Methods Seventy-eight patients with pancreatic carcinoma and 49 patients with pancreatic benign diseases were collected from January 2013 to December 2015.All the patients underwent EUS-FNA and cell or tissue samples were collected.DNA was extracted from the samples,and the codon 12 and 13 mutation of K-ras gene was detected by specific nano capture probe.Liquid cytology was also conducted.The sensitivity and specificity of the two methods were compared.Results The K-ras mutation rate was 92.3％ (72/78) in 78 patients with pancreatic carcinoma,which was obviously higher than the mutation rate of 20.4％ (10/49) in 49 patients with pancreatic benign tumors,and the difference was statistically significant (x2 =68.002,P =0.000).The sensitivity of the cytology examination of EUS-FNA specimens in the diagnosis of pancreatic carcinoma was 75.6％,the specificity was 87.8％,Youden index was 0.634,the positive and negative predictive value was 6.196 and 0.227.The detection of K-ras mutations had a sensitivity of 92.3％ and a specificity of 79.6％ in the diagnosis of pancreatic carcinoma,the Youden index was 0.719,the positive and negative predictive value was 4.524 and 0.096.K-ras mutation detection had a higher sensitivity compared with cytology,and the difference was statistically significant (x2 =8.47,P =0.004).The sensitivity was 94.9％ and specificity of 95.9％ using the combination of K-ras mutations and cytology for diagnosing pancreatic cancer,and the specificity was obviously increased compared with only k ras mutation detection and the difference was statistically significant (x2 =6.13,P =0.013).Conclusions K-ras mutation detection of EUS-FNA specimen using nano capture probe system can significantly improve the sensitivity of diagnosing pancreatic cancer,and the sensitivity and specificity could be further improved when combined with cytology.

Objective To explore the effect of let?7 miRNA silencing on Kaposi′s sarcoma? associated herpesvirus ( KSHV) lytic replication and the underling mechanism. Methods The pEGFP?C2?let?7 sponge vector was transfected into BCBL?1 and 293T cells with Lipofectamine 2000 to silence the expression of let?7 miRNAs. Quantitative real?time PCR ( qRT?PCR) was used to quantify the expression of let?7 miRNAs, the transcriptional levels of mitogen?activated protein kinase kinase kinase kinase 4 ( MAP4K4) , cyclooxygenase?2 ( COX?2) and matrix metalloproteinase 13 ( MMP?13) , and the DNA copy numbers of KSHV open reading frame 50 ( ORF50) and open reading frame 72 ( ORF72) . Western blot was used to detect the total and phosphorylated protein levels of MAP4K4, COX?2, extracellular regulated protein kinases (ERK1/2), c?Jun N?terminal kinase (JNK) and p38 MAPK. Results The expression of let?7 miRNAs was dramatically decreased in the let?7 sponge transfected BCBL?1 and 293T cells compared with that in the vector?transfected cells ( P<0.05 for all) . The gene copy number and mRNA transcriptional level of KSHV ORF50 were significantly increased in the let?7 sponge transfected BCBL?1 cells compared with that in the vector?transfected cells ( 1. 00 ± 0. 10 vs. 2. 33 ± 0. 18 and 1. 08 ± 0. 48 vs 3. 22 ± 0. 27, respectively, P<0.001 for both) . The gene copy number and mRNA transcriptional level of KSHV ORF72 were also significantly increased in let?7 sponge transfected BCBL?1 cells compared with those in the vector?transfected cells(1.07±0.49 vs 1.67±0.45 and 1.01±0.19 vs 1.54±0.11, respectively, P<0.05 for both). Furthermore, the mRNA transcriptional levels of MAP4K4, COX?2 and MMP?13 were significantly increased in the let?7 sponge transfected BCBL?1 cells compared with those in the vector?transfected cells (1.00±0.05 vs 5.73±0.96, 1.00±0.05 vs 2.68±0.19, 1.00±0.02 vs 2.69±0.25, respectively, P<0.001 for all). Let?7 miRNAs silencing also increased the protein expression levels of MAP4K4, COX?2 and phospho?ERK1/2, while the phospho?JNK and phospho?p38 were not changed in the BCBL?1 and 293T cells. Conclusions Let?7 silencing may activate the replication of KSHV, possibly through up?regulating MAP4K4 and its downstream molecules COX?2, MMP?13, and phosphorylation of ERK1/2, finally results in the progression of Kaposi sarcoma.

Objective To explore the effect of let?7 miRNA silencing on Kaposi′s sarcoma? associated herpesvirus ( KSHV) lytic replication and the underling mechanism. Methods The pEGFP?C2?let?7 sponge vector was transfected into BCBL?1 and 293T cells with Lipofectamine 2000 to silence the expression of let?7 miRNAs. Quantitative real?time PCR ( qRT?PCR) was used to quantify the expression of let?7 miRNAs, the transcriptional levels of mitogen?activated protein kinase kinase kinase kinase 4 ( MAP4K4) , cyclooxygenase?2 ( COX?2) and matrix metalloproteinase 13 ( MMP?13) , and the DNA copy numbers of KSHV open reading frame 50 ( ORF50) and open reading frame 72 ( ORF72) . Western blot was used to detect the total and phosphorylated protein levels of MAP4K4, COX?2, extracellular regulated protein kinases (ERK1/2), c?Jun N?terminal kinase (JNK) and p38 MAPK. Results The expression of let?7 miRNAs was dramatically decreased in the let?7 sponge transfected BCBL?1 and 293T cells compared with that in the vector?transfected cells ( P<0.05 for all) . The gene copy number and mRNA transcriptional level of KSHV ORF50 were significantly increased in the let?7 sponge transfected BCBL?1 cells compared with that in the vector?transfected cells ( 1. 00 ± 0. 10 vs. 2. 33 ± 0. 18 and 1. 08 ± 0. 48 vs 3. 22 ± 0. 27, respectively, P<0.001 for both) . The gene copy number and mRNA transcriptional level of KSHV ORF72 were also significantly increased in let?7 sponge transfected BCBL?1 cells compared with those in the vector?transfected cells(1.07±0.49 vs 1.67±0.45 and 1.01±0.19 vs 1.54±0.11, respectively, P<0.05 for both). Furthermore, the mRNA transcriptional levels of MAP4K4, COX?2 and MMP?13 were significantly increased in the let?7 sponge transfected BCBL?1 cells compared with those in the vector?transfected cells (1.00±0.05 vs 5.73±0.96, 1.00±0.05 vs 2.68±0.19, 1.00±0.02 vs 2.69±0.25, respectively, P<0.001 for all). Let?7 miRNAs silencing also increased the protein expression levels of MAP4K4, COX?2 and phospho?ERK1/2, while the phospho?JNK and phospho?p38 were not changed in the BCBL?1 and 293T cells. Conclusions Let?7 silencing may activate the replication of KSHV, possibly through up?regulating MAP4K4 and its downstream molecules COX?2, MMP?13, and phosphorylation of ERK1/2, finally results in the progression of Kaposi sarcoma.

Objective:To assess the rationality of the maximum lesion diameter of 15 mm in prostate imaging reporting and data system(PI-RADS)as a criterion for upgrading a lesion from category 4 to 5 and improve it to enhance the prediction of clinically significant prostate cancer(csPCa).Methods:In this study,the patients who underwent prostate magnetic resonance imaging(MRI)and prostate biopsy at Peking University First Hospital from 2019 to 2022 as a development cohort,and the patients in 2023 as a validation cohort were reviewed.The localization and maximum diameter of the lesion were fully evalua-ted.The area under the curve(AUC)and the cut-off value of the maximum diameter of the lesion to pre-dict the detection of csPCa were calculated from the receiver operating characteristics(ROC)curve.Confounding factors were reduced by propensity score matching(PSM).Diagnostic efficacy was com-pared in the validation cohort.Results:Of the 589 patients in the development cohort,358(60.8％)lesions were located in the peripheral zone and 231(39.2％)were located in the transition zone,and 496(84.2％)patients detected csPCa.The median diameter of the lesions in the peripheral zone was smaller than that in the transition zone(14 mm vs.19 mm,P＜0.001).In the ROC analysis of the maximal diameter on the csPCa prediction,there was no statistically significant difference between the peri-pheral zone(AUC=0.709)and the transition zone(AUC=0.673,P=0.585),and the cut-off values were calculated to be 11.5 mm for the peripheral zone and 16.5 mm for the migrating zone.By calcula-ting the Youden index for the cut-off values in the validation cohort,we found that the categorisation by lesion location led to better predictive results.Finally,the net reclassification index(NRI)was 0.170.Conclusion:15 mm as a criterion for upgrading the PI-RADS score from 4 to 5 is reasonable but too general.The cut-off value for peripheral zone lesions is smaller than that in transitional zone.In the future consideration could be given to setting separate cut-off values for lesions in different locations.