To study the role of BK virus(BK polyomavirus)in the development of the hemorrhagic cystitis(HC)after hematopoietic stem cell transplantation(HSCT)and analyze the risk fators for BK viruri4a and HC.Methods From August 2006 to November 2007,blood and urine samples were collected from 80 patients undergoing HSCT.BK virus DNA was detected with PCR.Cytomegalovirus (CMV)antigen was detected with immunofluorescence histochemical examination.A control group including 20 healthy individuals was established.Results Late-onset HC occurred in 15 of the 80 HSCT patients with an incidence of 18.8%.The median onset time of HC was 44(13-150)days after transplantation.BK viruria was detected in 30 of the 80 HSCT patients(37.5%)and the positive rate of viruria in the HC patients was 86.7%(13/15).The median time of BK viruria detection in HC patients wag 23(0-56)days after transplantation,being earlier than the onset time of HC.The persistence time of BK viruria was 7(2-14) weeks,being much longer than that of HC(11 days).CMV antigen viremia was detected in 12 of the 80 transplanted patients.with a positive rate of 36.7% in patients with BK viruria and 40.0% in HC patients.Nine of the 30 HC patients developed acute graft versus host disease(Agvhd)of grade Ⅱ-Ⅳ(30.0%).BK virus was not detected in the urine of the remainimg two HC patients and the 20 control subjects as well as in all the blood samples.Univariate analysis indicated that CMV viremia and Agvhd of grade Ⅱ-Ⅳ were agsociated with the occurrence of BK viruria.Condusions BK viruria is the main cauge of the late-onset HC after HSCT.CMV infection and Agvhd may contribute to the occurrence of HC agsociatieg with BK virus.

Objective To explore the non-HLA gene polymorphisms that influence CMV infection after hematopoietic stem cell transplantation (HSCT).Method Non-HLA gene (ACE,CD14,MPO,MBL) single nucleotide polymorphisms were determined by using sequence-specific primer polymerase chain reaction (PCR-SSP) and sequencing in 64 pairs of donors and recipients before HSCT and the differences of non-HLA gene were analysed in CMV positive and negative patients Results The distribution of ACE gene single nucleotide polymorphism was DD (14/128,10.9％),ID (72/128,56.3％),and Ⅱ (42/128,38.8％).The distribution of CD14-159 allele gene single nucleotide polymorphism was CC (18/128,14.1％),CT (81/128,63.3％),and TT (29/128,22.7％).The distribution of MPO-463 allele gene single nucleotide polyrnorphism was G (100)/128,78.1％),A (2/128,1.6％),and GA (26/128,20.3％).The distribution of MBL gene single nucleotide polymorphism was H (28/128,21.9％),HL (73/128,57.0％),L (27/128,21.1％),Y (87/128,68.0％),YX (38/128,29.7％),X (3/128,2.3％),A (94/128,73.4％),AB (32/ 128,25.0％),and B (2/128,1.6％).The allele frequency of ACE,CD14 and MPO shoed no significant differcence between CMV positive and negative patients The gene frequency of MBL-HL was increased in CMV positive group.Conclusion MBL gene single nucleotide polymorphisrns may influence CMV infection after HSCT.

Objective To investigate the frequency and mutational status of JAK2V617F mutation in Chinese patients with chronic myeloproliferative disorders(CMPD) and to study the relative quantification of mutated JAK2 mRNA and the clinical significance. Methods JAK2V617F mutation and the mutational status were screened with amplification-refractory mutation system polymerase chain reaction(ARMS-PCR), the relative quantification of mutated JAK2 mRNA was studied by using capillary electrophoresis. Results A higher prevalence of JAK2V617F in either the heterozygotc or homozyote status in essential thrombocythemia (ET) was observed in elderly patients with ET (P<0.05). The presence of JAK2V617F was found to be significantly correlated with the age at diagnosis (P<0.05); patients with age ≥ 60 years showed significantly higher JAK2 mutated RNA levels than those with age < 60 years (P<0.05); the presence of JAK2V617F in polycythemia vera (PV) and ET was found to be significantly associated with higher hemoglobin level and higher leukocyte count (P< 0.05). In addition, higher leukocyte count was observed in homozygous ET patients than in heterozygous ET patients (P<0.05). The frequency of JAK2V617F mutation and the prevalence of homozygote in PV patients were higher than those in ET patients (P<0.05). The differences of JAK2V617F mRNA levels among PV, ET and chronic idiopathic myelofibrosis (IMF) were not significant. Conclusions ARMS-PCR technique can be used to detect the frequency and mutational status of JAK2V617F mutation owing to its sensitivity and along with capillary electrophoresis, quantitative assay for mutated JAK2 mRNA, diagnosis of CMPD and judgement of prognosis become possible.

Objective To analyze the routine test parameter levels of patients with colorectal adenoma and colorectal cancer, and develop a prediction model. Methods A total of 580 patients diagnosed with colorectal adenoma (117 patients) and colorectal cancer (463 patients) were included in the retrospective study. The patients were randomly divided into two groups according to a 7:3 ratio: a training set with 406 cases and a validation set with 174 cases. Logistic regression analysis was used to establish a prediction model, and a nomogram was drawn. The model′s discrimination, calibration, and clinical applicability were evaluated using receiver operating characteristic curve (ROC), calibration plot, and decision curve analysis (DCA). Results Univariate logistic regression analysis identified 13 potential predictors: age, fecal occult blood test (FOBT), fibrinogen (FIB), thrombin time (TT), albumin (ALB), white blood cell value (WBC), neutrophil count (NEUT#), hematocrit value (HCT), mean corpuscular hemoglobin (MCH), red cell distribution width (RDW), platelet count (PLT), mean platelet volume (MPV), and activated partial thromboplastin time (APTT). Multivariate logistic regression analysis showed MPV, FIB, ALB, FOBT, TT, and HCT were risk factors for colorectal cancer in patients with colorectal adenoma (P<0.05). A nomogram was constructed based on these predictors to build a prediction model. The AUC of the ROC curve was 0.915 for colorectal cancer in the training set and 0.836 in the validation set. Calibration plots demonstrated high prediction accuracy and good model calibration. DCA results indicated the prediction model provided greater net benefit compared with the extreme models at threshold probabilities of approximately 55%-95%. Conclusion The developed prediction model exhibits satisfactory discrimination, calibration, and clinical applicability. The model can serve as an auxiliary tool in distinguishing between colorectal adenoma and colorectal cancer in patients.

Objective: To evaluate the non-carcinogenic health risk of heavy metals (As, Cd, Cr, Cu, Mn, Pb and Zn) in residential indoor dust for young children around an e-waste dismantling area in South China. Methods: A village around an e-waste dismantling area in South China was selected as a research site in October 2016. Convenience sampling method was used to select 36 houses in the village and 36 dust samples were collected by vacuum cleaner. The concentrations of heavy metals (Cd, Cr, Cu, Mn, Pb and Zn) in each sample were determined and expressed by the average value. Non-carcinogenic health risk assessment was conducted using the US Environmental Protection Agency (EPA) Health Risk Assessment (HRA) model, the American Toxicology and Disease Registry (ATSDR) Target-organ Toxicity Dose (TTD) approach and the ATSDR Binary Weight-of-Evidence (BINWOE) model. Results: The mean ± SD of concentrations of As, Cd, Cr, Cu, Mn, Pb and Zn were (48.90±33.91), (5.95±3.89), (173.57±580.37), (412.71±1 190.00), (612.82±540.70), (297.41±293.22) and (1 052.81±1 156.48) mg/kg, respectively. The HI value of TTD (2.670) and BINWOE (2.933) were higher than the safety threshold of EPA recommended non-carcinogenic health risk. The HI value of TTD and BINWOE were 1.93 and 2.12 times higher than the HI value of HRA (1.386). Conclusion There was non-carcinogenic health risk of heavy metals (As, Cd, Cr, Cu, Mn, Pb and Zn) via residential indoor dust around the e-waste dismantling area for local children.

Objective:To investigate the changes of various cytokines and coagulation function in B cell acute lymphoblastic leukemia(ALL) patients with different CRS scores during CD19-CAR-T cell immunotherapy.Methods:87 patients with B-ALL hospitalized in the First Affiliated Hospital of Soochow University and 30 normal controls were enrolled into this study from July 2018 to October 2020. The age of the patients was 32(20, 56) years old and 36(41.4%) were female. All these coagulation indicators, prothrombin time (PT), activated partial thromboplastin time (APTT), D-dimer, fibrinogen (Fg) were analyzed by automatic blood coagulation in B-ALL patients before and after treated with CAR-T cell. The ratio of CD19-CAR-T cells and the expression of IL-6, IL-10, IFN-γ, TFN-α, IL-2, IL-4, and IL-17A were analyzed using flow cytometry. The patients′ clinical parameters were detected, and the CRS classification of severity was made according to the standard of consensus.Results:Patients with CRS>3 had prolonged PT and APTT, increased D-dimer, and decreased fibrinogen ( P<0.05). The levels of cytokines of IFN-γ, IL-6, and IL-10 were significantly higher in patients with CRS>3 than that in controls ( P<0.05).The D-dimer level is positively correlated with IL-10. Conclusion:Patients with severe CRS grading have significant coagulation dysfunction in CD19-CAR-T cell immunotherapy. Cytokines IFN-γ, IL-6, and IL-10 may affect coagulation function and CRS grading during CD19-CAR-T cell immunotherapy.

ObjectiveTo observe the effect of the serum containing Puerariae Lobatae Radix on tumor necrosis factor-α （TNF-α）， endoplasmic reticulum stress signaling pathway protein endoplasmic reticulum stress protein glucose-regulated protein 78 （GRP78）， activated transcription factor 4 （ATF4）， protein kinase R-like endoplasmic reticulum kinase （PERK）， and eukaryotic translation initiation factors （eIF2α） and on inflammatory injury of macrophages induced by high glucose， and explore its possible mechanism. MethodThe control serum and serum containing Puerariae Lobatae Radix were prepared by serum pharmacology. The RAW264.7 macrophages were cultured in vitro， and 62.5 mmol·L^-1 glucose was used to induce macrophages to establish a model of severe endoplasmic reticulum （ER） stress （ERS） in vitro. Different volume fractions of serum containing Puerariae Lobatae Radix and ERS inhibitor （4-PBA） were used to interfere with the cells. Different glucose concentrations （22.5， 23.5， 25.0， 27.5， 32.5， 47.5， 72.5， 122.5 mmol·L^-1） and the effect of different volume fractions of serum containing Puerariae Lobatae Radix （0%， 2.5%， 5%， 7.5%， 10%， 15%， 20%） on the survival rate of RAW264.7 macrophages were detected by cell proliferation and cell counting kit-8 （CCK-8） assay. Based on the results of the effect of glucose concentrations on macrophage survival rate by CCK-8， Western blot （WB） was used to determine the protein expression levels of GRP78， the signature protein of ERS， by different concentrations of glucose （22.5， 32.5， 42.5， 52.5， 62.5， 72.5 mmol·L^-1） at different time periods （6， 12， 24， 36， 48 h）， and the optimal concentration and time for establishing the model of severe ERS were screened out. Based on the above experimental results， the cells were divided into blank group， 62.5 mmol·L^-1 glucose model group， 2 mmol·L^-1 4-PBA group， and high， medium， and low-dose （15%， 10%， 5%） serum containing Puerariae Lobatae Radix groups for subsequent experiments. The expression of TNF-α in the supernatant of RAW264.7 macrophages in the model of severe ERS was determined by enzyme-linked immunosorbent assay （ELISA）. The expressions of related pathway proteins in the model of severe ERS were determined by WB. ResultThe results of CCK-8 assay showed that the survival rate of macrophages reached the highest under the stimulation of glucose concentration of 27.5 mmol·L^-1 （P<0.01）， while the survival rate of macrophages increased with the concentration increasing from 22.5 mmol·L^-1 to 27.5 mmol·L^-1. When the glucose concentration was 25.0 mmol·L^-1， there was a significant difference （P<0.01）， and when the glucose concentration was 37.5 mmol·L^-1 to 122.5 mmol·L^-1， there was a downward trend. The serum containing Puerariae Lobatae Radix showed significant differences in the volume of 1% to 15% （P<0.05， P<0.01）. The results of WB found that the GRP78 protein expression was the most significant at 24 h （P<0.01）， and the GRP78 protein expression was the most significant when the glucose concentration was 62.5 mmol·L^-1 （P<0.01）. Therefore， 62.5 mmol·L^-1 of glucose was the optimal concentration to induce the model of severe ERS. As compared with the blank group， the protein expression levels of TNF-α， GRP78， ATF4， phosphorylation（p）-PERK/PERK， p-eIF2α/eIF2α in the model group were significantly increased （P<0.05， P<0.01）， and as compared with the model group， the protein expression levels of TNF-α， GRP78， ATF4， p-PERK/PERK， p-eIF2α/eIF2α in each administration group were significantly decreased （P<0.05， P<0.01）. ConclusionThe results of in vitro experiments show that Puerariae Lobatae Radix can alleviate the inflammatory injury of macrophages induced by high glucose to a certain extent and restore cell homeostasis by inhibiting the expression of GRP78， ATF4， PERK， and eIF2α in the ERS signaling pathway.

Proteins are the physical basis of life and perform all kinds of life activities. Proteins have different orientations and function in different tissues. The same protein, located in different subcellular regions, can perform different and even opposite functions. Both functional and structural proteins are capable of undergoing re-localization which can directly or indirectly participate in signal transduction. Due to abnormal transduction of signals during carcinogenesis, the proteins originally expressed in the cytoplasm are translocated into the nucleus and lead to functional changes in the tumor tissue. The changes of protein localization are affected by many factors, including the interaction between proteins, expression level of proteins and the cleaved intracellular domain of transmembrane protein.
Carcinogenesis ; pathology ; Cell Line, Tumor ; Cell Nucleus ; metabolism ; Cytoplasm ; metabolism ; Gene Expression Regulation, Neoplastic ; Humans ; Membrane Proteins ; metabolism ; Protein Domains ; Protein Transport ; physiology ; Signal Transduction

Objective: To investigate herpesvirus infection in early stage of hematopoietic stem cell transplantation (HSCT) by multiplex polymerase chain reaction (PCR), and to explore the association between multiple herpesviruses infection and clinical characteristics in HSCT patients and its impact on post-transplant complications and prognosis. Methods: A total of 734 peripheral blood samples were collected from 90 patients undergoing HSCT in the Department of Hematology, the First Affiliated Hospital of Soochow University between February 2017 and August 2017. The peripheral blood specimens were obtained before and within 90 days after transplantation at different time points. Lab-Aid824 Nucleic Acid Extraction Mini Reagent was used to extract DNA and multiplex PCR assay was used to simultaneously detect 8 kinds of human herpesviruses from genomic DNA. The incidence of various herpesvirus infections, its correlation with clinical features and effects on post-transplant complications and prognosis were analyzed. Results: The median follow-up time was 192 (range: 35-308) days. Among the 90 patients before transplantation, the incidence of herpes virus infection was 35.6% (32/90), including 12.2% (11/90) with one herpes virus infection and 23.3% (21/90) with multiple viruses infection. The incidence of herpes virus infection after transplantation was 77.8% (70/90), including 20.0% (18/90) with one herpes virus infection and 57.8% (52/90) with multiple herpes virus infection. Among the 52 patients with multiple herpes viruses infection, 30 (57.7%) patients were infected by 2 kinds of viruses, 18 (34.6%) patients by 3 kinds of viruses and 4 (7.7%) patients by 4 kinds of viruses. There was a correlation between HHV-6 and HHV-7 herpesvirus infection (OR=13.880, Q=0.026). EBV infection was related to HHV-7 infection (OR=0.093, Q=0.044). The age of patients was correlated with the incidence of HHV-1 infection before transplantation. There were 24 patients in our study experienced clinical symptoms associated with viral infection. The main manifestations were hemorrhagic cystitis (HC), interstitial pneumonia, enteritis, viral encephalitis and fever of unknown origin. EBV infection was related to HLA incompatibility and the inconsistent of the ABO blood group and grade Ⅱ-Ⅳ aGVHD after transplantation. HLA incompatibility and the unrelated donor and grade Ⅱ-Ⅳ aGVHD were related to multiple viruses infection. Conclusion Multiple herpesviruses were common in patients undergoing HSCT, which were closely related to HLA mismatch, unrelated donor and grade Ⅱ-Ⅳ aGVHD.

ObjectiveTo explore the mechanism of Buyang Huanwutang in regulating macrophage polarization based on the Toll-like receptor 4 （TLR4） / nuclear factor-κB （NF-κB） / nucleotide-binding oligomerization domain-like receptor 3 （NLRP3） pathway. MethodRAW264.7 macrophages were intervened with lipopolysaccharide （LPS） of different concentrations （0， 1.25， 2.5， 5， 10， 20， 40， and 80 mg·L^-1） for 24 hours. Cell Counting Kit-8 （CCK-8） assay was used to determine the cell viability of RAW264.7 macrophages. The optimal concentration was chosen to establish an in vitro inflammation model induced by LPS. Cells were divided into a blank group （20% blank serum）， a model group （20% blank serum + 10 mg·L^-1 LPS）， a model control group （20% FBS + 10 mg·L^-1 LPS）， low-， medium-， and high-dose （5%， 10%， and 20%） Buyang Huanwutang-containing serum groups， a high-dose （20%） Buyang Huanwutang combined with NLRP3 inhibitor MCC950 （50 μmol·L^-1） group， a high-dose （20%） Buyang Huanwutang combined with reactive oxygen species （ROS） inhibitor NAC （10 μmol·L^-1） group， and a high-dose （20%） Buyang Huanwutang combined with NF-κB inhibitor PDTC （10 μmol·L^-1） group. Enzyme-linked immunosorbent assay （ELISA） was used to detect the expression of interleukin-1β （IL-1β）， interleukin-18 （IL-18）， and tumor necrosis factor-α （TNF-α） in RAW264.7 macrophages. Flow cytometry was employed to measure ROS levels in macrophages. Western blot was used to determine the protein expression of M1-type macrophage-related factors inducible nitric oxide synthase （iNOS） and TNF-α， M2-type macrophage-related factors arginase-1 （Arg-1） and interleukin-10 （IL-10）， as well as the proteins in the TLR4/NF-κB/NLRP3 pathway. ResultCCK-8 results indicated that under 10 mg·L^-1 LPS stimulation， RAW264.7 macrophages exhibited the highest cell viability （P<0.01）. Compared with the blank group， the model group showed significantly increased levels of IL-1β， IL-18， and TNF-α （P<0.05，P<0.01）， increased ROS expression （P<0.05，P<0.01）， increased protein expression of M1-type macrophage factors iNOS and TNF-α （P<0.01）， decreased protein expression of M2-type macrophage factors Arg-1 and IL-10 （P<0.05，P<0.01）， and upregulated expression levels of TLR4， myeloid differentiation factor 88 （MyD88）， phosphorylated inhibitor of NF-κB （p-IκB）/NF-κB inhibitor （IκB）， phosphorylated NF-κB （p-NF-κB） p65/NF-κB p65， NLRP3， apoptosis-associated speck-like protein containing a CARD （ASC）， and pro-Caspase-1 （P<0.05， P<0.01）. Compared with the model group， all Buyang Huanwutang-treated groups and inhibitor groups significantly reduced levels of IL-1β， IL-18， and TNF-α （P<0.01）， suppressed the expression of inflammatory factors in RAW264.7 macrophages， decreased cellular ROS expression levels （P<0.01）， downregulated M1-type macrophages iNOS and TNF-α protein expression （P<0.01）， upregulated M2-type macrophages Arg-1 and IL-10 protein expression （P<0.01）， and lowered protein expression levels of TLR4， MyD88， p-IκB/IκB， p-NF-κB p65/NF-κB p65， NLRP3， ASC， and pro-Caspase-1 （P<0.05， P<0.01）. ConclusionBuyang Huanwutang can improve macrophage inflammation， potentially by reducing macrophage ROS levels， inhibiting RAW264.7 macrophage polarization， and downregulating the protein expression levels of the TLR4/NF-κB/NLRP3 pathway.