Improving the Quality of Low-grade Tobacco by Enzymatic Treatment and Co-fermentation with Yeast and Lactic Acid Bacteria.

Enzymatic treatment is a promising method to modulate the chemical composition, flavoring substances and to enhance the sensory quality of cigarettes. This study investigates the feasibility of enzymatic treatment in conjunction with co-fermentation with yeast and lactic acid bacteria to improve the quality of low-quality cigarettes. Amylase, flavourzyme, glucoamylase, protease, and their combinations were used for the enzymatic treatment of tobacco-sorghum by using the liquid-state fermentation method. The biochemical components and flavor substances of the fermented products were analyzed. The findings show that amylase and glucoamylase can effectively degrade starch into fermentable reducing sugars, facilitate microbial growth and proliferation, and significantly enhance the levels of flavoring alcohols and esters. Flavourzyme and glucoamylase, either individually or combined with amylase, achieved a more balanced distribution of flavor substances in the products. Additionally, flavourzyme was capable of increasing the content of guaiacol, solanesol, and 2-acetylpyrrole in the fermented products, thereby improving the richness and depth of the cigarette flavor. While the protease treatment group showed fewer flavor substances and higher nicotine content, which was detrimental to the quality of cigarettes. When integrating the fermentation products into low-quality cigarettes, flavourzyme and glucoamylase combined with amylase treatment received higher scores in sensory quality evaluations. This study provides a beneficial strategy for effectively improving the quality of low-quality cigarettes.

Isotope dilution-HPLC-MS/MS to investigate the production patterns and possible pathways of free and protein-bound AGEs and 4-MI in cookies.

A qualitative and quantitative method for detecting free and protein-bound advanced glycation end products (AGEs) and 4-methylimidazole (4-MI) was established using isotope dilution-HPLC-MS/MS, and successfully applied in cookies and model systems. The effects of different temperatures (160-220 °C) on the formation of free and protein-bound AGEs and 4-MI in cookies were discussed, and the possible model systems (Maillard reaction pathway 1 using wheat gluten protein + glucose + sucrose; direct addition pathway 1 using wheat gluten protein + CML/CEL/4-MI) of protein-bound AGEs and 4-MI were verified. The results showed that the contents of protein-bound CML, CEL, and 4-MI were higher than free content with a tendency of increasing first and subsequently decreasing with temperature, reaching a maximum at 200 °C in cookies. In the model systems, the levels of protein-bound CML, CEL, and 4-MI are higher than those of free CML, CEL, and 4-MI. The protein-bound CML, CEL, and 4-MI accounted for 90.73, 87.64, and 97.56% of the total amount in the model system 1, while accounting for 68.19, 59.00, and 50.96% in the model system 2, respectively. In comparison, protein-bound CML, CEL, and 4-MI could be easily generated directly by Maillard reaction.

Wilms' tumor 1-associating protein plays an aggressive role in diffuse large B-cell lymphoma and forms a complex with BCL6 via Hsp90.

BACKGROUND: Wilms' tumor 1-associating protein (WTAP) is a nuclear protein, which is ubiquitously expressed in many tissues. Furthermore, in various types of malignancies WTAP is overexpressed and plays a role as an oncogene. The function of WTAP in diffuse large B-cell lymphoma (DLBCL), however, remains unclear. METHODS: Immunohistochemistry was applied to evaluate the levels of WTAP expression in DLBCL tissues and normal lymphoid tissues. Overexpression and knock-down of WTAP in DLBCL cell lines, verified on mRNA and protein level served to analyze cell proliferation and apoptosis in DLBCL cell lines by flow cytometry. Finally, co-immunoprecipitation (Co-IP), IP, and GST-pull down assessed the interaction of WTAP with Heat shock protein 90 (Hsp90) and B-cell lymphoma 6 (BCL6) as well as determined the extend of its ubiquitinylation. RESULTS: WTAP protein levels were consistently upregulated in DLBCL tissues. WTAP promoted DLBCL cell proliferation and improved the ability to confront apoptosis, while knockdown of WTAP in DLBCL cell lines allowed a significant higher apoptosis rate after treatment with Etoposide, an anti-tumor drug. The stable expression of WTAP was depended on Hsp90. In line, we demonstrated that WTAP could form a complex with BCL6 via Hsp90 in vivo and in vitro. CONCLUSION: WTAP is highly expressed in DLBCL, promoting growth and anti-apoptosis in DLBCL cell lines. WTAP is a client protein of Hsp90 and can appear in a complex with BCL6 and Hsp90 in DLBCL. Down-regulation of WTAP could improve the chemotherapeutic treatments in DLBCL.

LITAF is a potential tumor suppressor in pancreatic cancer.

Early diagnosis of pancreatic cancer, one of the most deadly cancers with low survival rates, is difficult, and effective biomarkers are urgently needed. Lipopolysaccharide-induced tumor necrosis factor-α factor (LITAF) has been recently proposed as a potential tumor suppressor gene in several types of cancer. Here, we analyzed the biological function of LITAF in pancreatic cancer. The LITAF gene and protein levels were decreased in pancreatic tumor tissues compared with their paired adjacent non-cancerous tissues. In addition, patients with the lower LITAF protein expression had lower disease-free survival rates. The decreased LITAF expression correlated with LITAF promoter hypermethylation in pancreatic cancer cells and tissues. Moreover, promoter demethylation dose-dependently increased the LITAF transcription. Importantly, LITAF demethylation suppressed proliferation and cell cycle progression, and enhanced apoptosis of pancreatic cancer cells. Together, our results indicate that LITAF functions as a tumor suppressor gene in pancreatic cancer cells, and might serve as a novel biomarker for early diagnosis of pancreatic cancer.

The feedback loop of LITAF and BCL6 is involved in regulating apoptosis in B cell non-Hodgkin's-lymphoma.

Dysregulation of the apoptotic pathway is widely recognized as a key step in lymphomagenesis. Notably, LITAF was initially identified as a p53-inducible gene, subsequently implicated as a tumor suppressor. Our previous study also showed LITAF to be methylated in 89.5% B-NHL samples. Conversely, deregulated expression of BCL6 is a pathogenic event in many lymphomas. Interestingly, our study found an oppositional expression of LITAF and BCL6 in B-NHL. In addition, LITAF was recently identified as a novel target gene of BCL6. Therefore, we sought to explore the feedback loop between LITAF and BCL6 in B-NHL. Here, our data for the first time show that LITAF can repress expression of BCL6 by binding to Region A (-87 to +65) containing a putative LITAF-binding motif (CTCCC) within the BCL6 promoter. Furthermore, the regulation of BCL6 targets ( PRDM1 or c-Myc) by LITAF may be associated with B-cell differentiation. Results also demonstrate that ectopic expression of LITAF induces cell apoptosis, activated by releasing cytochrome c, cleaving PARP and caspase 3 in B-NHL cells whereas knockdown of LITAF robustly protected cells from apoptosis. Interestingly, BCL6, in turn, could reverse cell apoptosis mediated by LITAF. Collectively, our findings provide a novel apoptotic regulatory pathway in which LITAF, as a transcription factor, inhibits the expression of BCL6, which leads to activation of the intrinsic mitochondrial pathway and tumor apoptosis. Our study is expected to provide a possible biomarker as well as a target for clinical therapies to promote tumor cell apoptosis.

The truncate mutation of Notch2 enhances cell proliferation through activating the NF-κB signal pathway in the diffuse large B-cell lymphomas.

The Notch2 is a critical membrane receptor for B-cell functions, and also displays various biological roles in lymphoma pathogenesis. In this article, we reported that 3 of 69 (4.3%) diffuse large B-cell lymphomas (DLBCLs) exhibited a truncate NOTCH2 mutation at the nucleotide 7605 (G/A) in the cDNA sequence, which led to partial deletion of the C-terminal of PEST (proline-, glutamic acid-, serine- and threonine-rich) domain. The truncate Notch2 activated both the Notch2 and the NF-κB signals and promoted the proliferation of B-cell lymphoma cell lines, including DLBCL and Burkitt's lymphoma cell lines. Moreover, the ectopic proliferation was completely inhibited by ammonium pyrrolidinedithiocarbamate (PDTC), an NF-κB inhibitor. Simultaneously, PDTC also reduced the expression level of Notch2. Based on these results, we conclude that the Notch2 receptor with PEST domain truncation enhances cell proliferation which may be associated with the activation of the Notch2 and the NF-κB signaling. Our results are expected to provide a possible target for new DLBCL therapies by suppressing the Notch2 and the NF-κB signaling.

Critical role of SHP2 (PTPN11) signaling in germinal center-derived lymphoma.

Germinal center lymphoma is a heterogeneous human lymphoma entity. Here we report that constitutive activity of SHP2 (PTPN11) and its downstream kinase ERK is essential for the viability of germinal center lymphoma cells and disease progression. Mechanistically, SHP2/ERK inhibition impedes c-Myc transcriptional activity, which results in the repression of proliferative phenotype signatures of germinal center lymphoma. Furthermore, SHP2/ERK signaling is required to maintain the CD19/c-Myc loop, which preferentially promotes survival of a distinct subtype of germinal center lymphoma cells carrying the MYC/IGH translocation. These findings demonstrate a critical function for SHP2/ERK signaling upstream of c-Myc in germinal center lymphoma cells and provide a rationale for targeting SHP2 in the therapy of germinal center lymphoma.

Evaluation of specific marker CK13 and CK10/13 combined with APM staining for the diagnosis of amniotic fluid embolism and aspiration.

OBJECTIVE: To explore the value of CK13 (Rab) and CK10/13(Mab) as objective and specific biomarkers combined with Alcian-Phloxine-Martius yellow (APM) staining for the diagnosis of amniotic fluid embolism (AFE) and amniotic fluid aspiration (AFA). METHODS: A retrospective study of forensic autopsy cases of 148 neonatal deaths and 19 maternal deaths in the perinatal stage was conducted at the Institute of Pathology and Forensic Medicine, Zhejiang University. Medical records were reviewed and monoclonal antibody for CK13 (Rab) and CK10/13 (Mab) as immunostaining of amniotic fluid squamous cells, APM staining, and Hematoxylin and Eosin (HE) staining were used to diagnose the AFE and AFA. Descriptive statistics of the patient population were analyzed using SPSS 20.0 software. RESULTS: Immunoreactivity of CK13 and CK10/13 specifically identified squamous cells of all the AFE and AFA cases. The amniotic fluid squamous cells were stained positive in a deep brown color with the monoclonal antibody to CK 13 and CK10/13 whereas the endothelial cells and alveolar epithelial cells were negative. There was no CK13 or CK10/13 expression in the non-AFE and non-AFA cases. With APM staining keratinized squamous cells were pink and mucus was blue-green in marked contrast with the surrounding tissue, which improved the detection rates of both keratinized squamous cells and mucus. CONCLUSIONS: CK13 (Rab) and CK10/13 (Mab) are valuable and reliable biomarkers of amniotic fluid squamous cells. APM reveals enriched mucus and keratinized squamous cells of amniotic fluid. Immunohistochemical detection of CK13 and CK10/13 combined with APM staining can improve the accuracy and reduce the difficulty in the diagnosis of AFE and AFA.