<div>Abstract<p><b>Purpose:</b> Hans and coworkers previously developed an immunohistochemical algorithm with ∼80% concordance with the gene expression profiling (GEP) classification of diffuse large B-cell lymphoma (DLBCL) into the germinal center B-cell–like (GCB) and activated B-cell–like (ABC) subtypes. Since then, new antibodies specific to germinal center B-cells have been developed, which might improve the performance of an immunostain algorithm.</p><p><b>Experimental Design:</b> We studied 84 cases of cyclophosphamide-doxorubicin-vincristine-prednisone (CHOP)–treated DLBCL (47 GCB, 37 ABC) with GCET1, CD10, BCL6, MUM1, FOXP1, BCL2, MTA3, and cyclin D2 immunostains, and compared different combinations of the immunostaining results with the GEP classification. A perturbation analysis was also applied to eliminate the possible effects of interobserver or intraobserver variations. A separate set of 63 DLBCL cases treated with rituximab plus CHOP (37 GCB, 26 ABC) was used to validate the new algorithm.</p><p><b>Results:</b> A new algorithm using GCET1, CD10, BCL6, MUM1, and FOXP1 was derived that closely approximated the GEP classification with 93% concordance. Perturbation analysis indicated that the algorithm was robust within the range of observer variance. The new algorithm predicted 3-year overall survival of the validation set [GCB (87%) versus ABC (44%); <i>P</i> < 0.001], simulating the predictive power of the GEP classification. For a group of seven primary mediastinal large B-cell lymphoma, the new algorithm is a better prognostic classifier (all “GCB”) than the Hans' algorithm (two GCB, five non-GCB).</p><p><b>Conclusion:</b> Our new algorithm is significantly more accurate than the Hans' algorithm and will facilitate risk stratification of DLBCL patients and future DLBCL research using archival materials. (Clin Cancer Res 2009;15(17):5494–502)</p></div>
Abstract Histamine alters the Th1/Th2 cytokine balance towards the Th2 cytokine profile and consequently play a role in allergic disease and asthma. This study was designed to determine if the effects of histamine on Signal Transducer and Activator of Transcription(STAT)-6 were direct or mediated via release of IL-4 production.The splenocytes from IL-4 knockout C57Bl/6 mice were pretreated with antibody to IL-13 for 30 minutes followed by treatment with histamine. The cells were then stimulated with PMA + ininomycin for 6 hours under standard cell culture conditions. For the control experiments splenocytes from the wild type C57BL/6 mice were used. The cells were lyzed and the total basal and phosphorylated STAT6 were analyzed using the Western Blot Analysis. Furthermore, splenocytes from C57Bl/6 mice were pretreated with antibodies to IL-4 and IL-13 followed by treatment with histamine and PMA + ionomycin and Western blot analysis were performed to determine the phosphorylated Stat6 levels.In the control group histamine caused hyper-phosphorylation of Y641 tyrosine residues of STAT6. These effects were not observed when splenocytes from the IL-4 knockout mice were used or the wildtype splenocytes were pretreated with antibodies to IL-4 and IL-13. These observations suggest that histamine indirectly affected the heper-phosphorylation of STAT6 via its effects on the secretion of IL-4 and H1 receptors played a role in this process, since H1 receptor antagonists blocked these effects. The clinical significance of STAT6 hyper-phosphorylation in the exacerbation of asthma symptoms has not yet been determined.
We aimed to quantify the extent of salivary gland fibrosis using shear-wave elastography (SWE) to assess its diagnostic value for primary Sjögren syndrome (pSS).A total of 58 pSS patients and 44 controls underwent SWE ultrasound evaluation of the parotid and submandibular glands. We measured the degree of salivary gland fibrosis in all participants and investigated the diagnostic accuracy of SWE for pSS and its relationship to disease progression.The diagnostic sensitivity, specificity, and accuracy of pSS were highest when the critical Young's modulus values of the parotid and submandibular glands were 18.4 and 15.9 kPa, respectively, effectively improving the diagnostic value of pSS. The area under the SWE curve of the submandibular gland was higher than that of the parotid gland (z = 2.292, P = 0.02), suggesting that the submandibular gland was damaged earlier. The mean parotid gland thickness of pSS patients was thicker than in healthy controls (mean ± standard deviation 2.5 ± 0.3 vs 2.4 ± 0.2, P = 0.013]. SWE had a 70.3% sensitivity for diagnosing pSS patients with a disease duration of 5 years, but this did not differ significantly from pSS patients with a longer disease duration.SWE is a valid diagnostic method for pSS. The degree of salivary gland fibrosis related to secretory function and pathological progression, and quantitative measurements of tissue elasticity provide objective criteria for predicting damage in pSS.
Lmx1a plays a central role in the specification of dopaminergic (DA) neurons, which potentially could be employed as a key factor for trans-differentiation to DA neurons. In our previous study, we have converted somatic cells directly into neural stem cell—like cells, namely induced neural stem cells (iNSCs), which further can be differentiated into subtypes of neurons and glia in vitro. In the present study, we continued to test whether these iNSCs have therapeutic effects when transplanted into a mouse model of Parkinson's disease (PD), especially when Lmx1a was introduced into these iNSCs under a Nestin enhancer. iNSCs that over-expressed Lmx1a (iNSC-Lmx1a) gave rise to an increased yield of dopaminergic neurons and secreted a higher level of dopamine in vitro. When transplanted into mouse models of PD, both groups of mice showed decreased ipsilateral rotations; yet mice that received iNSC-Lmx1a vs. iNSC-GFP exhibited better recovery. Although few iNSCs survived 11 weeks after transplantation, the improved motor performance in iNSC-Lmx1a group did correlate with a greater tyrosine hydroxylase (TH) signal abundance in the lesioned area of striatum, suggesting that iNSCs may have worked through a non-autonomous manner to enhance the functions of remaining endogenous dopaminergic neurons in brain.
CD19 chimeric antigen receptor T cell (CD19CAR-T) has shown great potential to treat acute B cell lymphoblastic leukemia (B-ALL) and B cell lymphoma, and most of anti-CD19 scFv are derived from murine antibody sequences. However, about 10-20% of B-ALL patients exhibit primary resistance to murine-based CD19CAR-T (CD19mCAR-T). Herein, we report that a humanized selective CD19CAR-T (CD19hsCAR-T) may offer a solution to this problem.
The use of small interfering RNAs (siRNAs) to silence target gene expression has greatly facilitated mammalian genetic analysis by generating loss-of-function mutants. In recent years, high-throughput, genome-wide screening of siRNA libraries has emerged as a viable approach. Two different methods have been used to generate short hairpin RNA (shRNA) libraries; one is to use chemically synthesized oligonucleotides, and the other is to convert complementary DNAs (cDNAs) into shRNA cassettes enzymatically. The high cost of chemical synthesis and the low efficiency of the enzymatic approach have hampered the widespread use of screening with shRNA libraries.We report here an improved method for constructing genome-wide shRNA libraries enzymatically. The method includes steps of cDNA fragmentation and endonuclease MmeI digestion to generate 19-bp fragments, capping the 19-bp cDNA fragments with a hairpin oligonucleotide, and amplification of the hairpin structures by PCR. The PCR step converts hairpins into double-stranded DNAs that contain head-to-head cDNA fragments that can be cloned into a vector downstream of a Pol III promoter.This method can readily be used to generate shRNA libraries from a small amount of mRNA and thus can be used to create cell- or tissue-specific libraries.
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