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Objective To investigate the expressions of programmed death-ligand 1 (PD-L1) and PD-L2 and phosphorylated protein kinase B (p-AKT) in diffuse large B-cell lymphoma (DLBCL) patients and their correlations with clinicopathological features and prognosis. Methods A total of 68 paraffin-embedded specimens of DLBCL patients diagnosed in Shanxi Provincial Cancer Hospital with detailed follow-up record from January 2010 to December 2012 were included in the study. The expressions of PD-L1, PD-L2 and p-AKT proteins in DLBCL were detected by using immunohistochemistry (IHC). Results The positive rate of PD-L1 protein in DLBCL patients was 22.1％ (15/68), which was related to germinal center B-cell (GCB) subtype or not (χ2= 5.591, P= 0.018), clinical stage (χ2= 3.969, P= 0.046), international prognostic index (IPI) grades (χ2=4.178, P=0.041) and treatment remission rate (χ2=6.587, P=0.010). The positive rate of PD-L2 protein in DLBCL patients was 14.7％ (10/68), which was related to extranodal metastasis or not (χ2=6.772, P= 0.009). The positive rate of p-AKT for DLBCL patients was 61.8％ (42/68), which was correlated with age (≥60 years old) or not (χ2=6.227, P=0.013), Eastern Cooperative Oncology Group (ECOG) grades (χ2=4.005, P=0.045), B symptoms (χ2=10.187, P=0.001) and treatment remission rate (χ2=4.096, P=0.043). Univariate survival analysis showed that the overall survival (OS) rate and progression free survival (PFS) rate of PD-L1 protein positive expression group were lower than those of PD-L1 protein negative expression group (both P< 0.05). In the patients with non-GCB subtype, OS rate and PFS rate of PD-L1 protein positive expression group were lower than those of PD-L1 protein negative expression group (both P<0.05). p-AKT protein positive expression group had poorer OS rate and PFS rate compared to p-AKT negative expression group (both P< 0.05). Correlation analysis showed that PD-L1 protein expression was correlated with PD-L2 and p-AKT proteins expressions (r= 0.380, P= 0.001;r= 0.273, P= 0.025). The prognosis was worse when p-AKT and PD-L1 proteins was co-expressed (P< 0.05). Multivariate analysis suggested high expressions of PD-L1 and p-AKT proteins were independent prognosis risk factors in DLBCL (both P<0.05). Conclusions The expressions of PD-L1 and p-AKT proteins may be involved in the occurrence and development of DLBCL. Blocking PD-1 and PD-L1 access or combined blocking could provide a promising future for the clinical therapy.
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Objective@#To optimize the preparation process for Haoqin-Huaban granules so as to provide experimental basis for the development and utilization of the compound.@*Methods@#Taking the extraction rate of gentiopicroside from Gentiana macrophylla Pall as the observation index, L9(34)orthogonal test was used to investigate the amount of water, extraction time, and extraction frequency for optimizing the water extracting technology of the Haoqin-Huaban granules. Then the dry paste was used for the raw material, the ratio of the diluent (dextrin) and the concentration and dosage of the binder (ethanol) were investigated by single factor, and the particles were prepared.@*Results@#The optimum water extraction process was A2B2C3, which indicated that the prescribed medicinal materials with 8 times water were extracted 1 h for 3 times. The extract was concentrated into paste, vacuum drying, pulverizing, mixing according to the ratio of 5:1, with ethanol, made of soft material, pelletizing, drying at 60 ℃ for granulation, in order to make Haoqin-Huaban granules.@*Conclusions@#The preparation process is reasonable and feasible, which provides experimental basis for the industrial production of Haoqin-Huaban granules.
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Objective To explore the relationship between myc/bcl-2 and myc/p53 co-expression and the prognosis of diffuse large B-cell lymphoma (DLBCL). Methods A total of 148 DLBCL cases with the follow-up data in Shanxi Cancer Hospital from January 2010 to October 2014 were selected. The expression of myc, bcl-2 and p53 protein in paraffin samples was detected by immunohistochemistry (IHC). Results The positive expression rate of myc, bcl-2 and p53 was 35.1 % (52/148), 60.1 % (89/148) and 24.3 % (36/148) respectively in 148 patients with DLBCL. There were 37 (25.0%) cases of myc/bcl-2 co-expression, which were correlated with Hans classification (χ2= 4.749, P= 0.029), IPI score (χ2= 4.894, P= 0.027), bone marrow invasion (χ2= 4.751, P= 0.029), and efficacy evaluation (χ2= 9.14, P= 0.003). Myc/p53 co-expression was detected in 17 cases (11.5 %), which was associated with Hans classification (χ2 = 5.349, P= 0.021) andLDH level (χ2= 11.1, P= 0.001). Univariate analysis revealed that myc [overall survival (OS): χ2= 6.044, P= 0.014; progression free survival (PFS):χ2= 6.212, P= 0.013], bcl-2 (OS:χ2= 5.812, P= 0.016; PFS:χ2= 4.878, P= 0.027), p53 (OS:χ2= 20.092, P< 0.0001; PFS:χ2= 18.492, P< 0.0001), myc/bcl-2 co-expression (OS: χ2= 11.277, P= 0.001; PFS:χ2= 9.024, P= 0.003) and myc/p53 co-expression (OS: χ2=21.150, P< 0.0001; PFS: χ2 = 18.655, P< 0.0001) were the adverse prognostic factors. In addition, the survival rate of the co-expression group was lower than that of single expression group, and the survival rate of myc/p53 co-expression group was lower than that of myc/bcl-2 co-expression group. Multivariate analysis showed that among the six independent variables, including myc, bcl-2, p53, myc/bcl-2, myc/p53 and treatment regimen, p53 expression was an independent prognostic affecting factor of OS (95%CI 0.172ˉ0.763, P=0.008) and PFS (95%CI 0.172ˉ0.773, P=0.009). Conclusion Myc, bcl-2 and p53 are the poor prognostic factors in DLBCL. Myc/bcl-2 and myc/p53 co-expression have synergistic effect, indicating the poor prognosis.
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Ulta performance liqiuid chromatography-triple quadrupole tandem mass spectrometry ( UPLC-MS/MS) was used to monitor the relative levels of bufadienolides in toad venom in normal and bensulfuron-polluted groups. Methanol extract of toad venom was separated by UPLC ( ODS-C18 ) using a gradient elution of water contains 0. 1% formic acid and acetonitrile. Mass spectrometry was used in an ESI source operated in positive ion and MRM mode. The parameters in the source were set as follows: capillary voltage 3. 0 kV; sampling cone voltage 30 V; and desolvation temperature 500℃. In this method, external calibrations of 6 standards were typically constructed (R2=0. 9953-0. 9992). The LOD was 0. 42-4. 86 ng/mL. Intra- and inter-day precision was 3. 8%-6. 8% and 4. 0%-8. 8%, respectively. The recovery of standard was evaluated by spiking the standard compound into toad venom. Their average recoveries were 96. 9%-109. 6%, and RSDs were 2. 0%-8. 1%. This method was further employed into monitoring the levels of 36 bufadienolides. The levels of more than 20 bufadienolides were greatly different after bensulfuron pollution, suggesting that the bensulfuron pollution could change the chemical expression pattern of bufadienolides in toad venom.