Esophageal and Gastric Varices ; etiology ; therapy ; Hemodynamics ; Humans ; Hypertension, Portal ; complications ; therapy ; Liver Cirrhosis ; complications ; therapy ; Portal Vein ; physiopathology

Objective To construct plasmids for knock-out of protein arginine methyltransferase 3 (prmt3) gene using CRISPR/Cas9 gene editing method and examine the effect of prmt3 knockout on the proliferation of human non-small cell lung cancer(NSCLC)A549 cells.Methods Synthesized sgRNA oligos targeting prmt3 gene were cloned into LentiCRISPR vector and positive constructs confirmed by sequencing later .After infection with the packaged virus , A549 cells were screened with puromycin , and then the single clones were isolated .The protein level of PRMT3 in individual cell clones was analyzed with Western blot . Biological assay of clone formation , wound healing , flow cytometry assay and mass spectrometry ( MS) analysis were used to compare cellular proliferation behavior changes between control cells and cells with prmt3 gene knockout .Results The LentiCRISPR plasmids targeting prmt3 gene were confirmed by sequencing , and the PRMT3 protein level was significantly decreased in PRMT 3 KO cells compared with control cells .Depletion of PRMT3 promoted cell proliferation and led to cell cycle arrest at G 2/M phase, but had no influence on cell migration .Besides, some PRMT3 substrate candidates were identified with mass spectrum assays .Conclusion A549 cells with prmt3 gene knockout based on CRISPR/Cas9 are successfully established .PRMT3 can regulate cell cycle and proliferation .

OBJECTIVETo explore the effect of perioperative nutrition support on nutritional condition and complications of the patients with postoperative pancreatic head cancer.

METHODSThirty four patients received perioperative nutrition support, including enteral nutrition and parenteral nutrition (treatment group). Forty eight patients received routine postoperative parenteral nutrition (control group). According to the operative method, these two groups were further divided into two sub-groups: (1) pancreaticoduodenectomy (PD) subgroup, including 13 cases from treatment group, and 24 cases from control group; (2) palliative operation subgroup, including 21 cases from treatment group, and 24 cases from control group. Body weight, total protein (TP), serum albumin (ALB), and the complications after operation were compared.

RESULTSThe concentrations of ALB and TP in the treatment group were significantly higher than those in the control group (P< 0.05). Body weight and TP of the patients received PD in the treatment group were significantly better than those of the control group (P < 0.05).

CONCLUSIONPerioperative nutrition support can improve postoperative nutritional condition and reduce the postoperative complications in patients with pancreatic head cancer.

Adult ; Aged ; Combined Modality Therapy ; Enteral Nutrition ; Female ; Humans ; Male ; Middle Aged ; Nutritional Support ; methods ; Pancreatic Neoplasms ; surgery ; therapy ; Pancreaticoduodenectomy ; Parenteral Nutrition ; Postoperative Complications ; prevention & control

In order to explore the application of real-time quantitative reverse-transcription polymerase chain reaction (Q-PCR) for detecting PML/RARalpha gene transcripts in patients with acute promyelocytic leukemia (APL), the bone marrow samples from 46 newly diagnosed APL patients were collected for analysis. Three plasmids containing cDNA fragments of the bcr1-, bcr3-form PML/RARalpha and ABL control gene were constructed respectively. The ABI Prism 7500 Sequence Detection System using Taqman fluorogenic probes was used to quantify target gene. PML/RARalpha mRNA was detected by Q-PCR in 46 APL patients and 40 non-APL patients. The normalized quotient (NQ) of PML/RARalpha mRNA was calculated as followings: NQ = PML/RARalpha mRNA copy numbers/ABL mRNA copy numbers. Immunophenotype of acute promyelocytic leukemia was determined by four-color flow cytometry. The results showed that the coefficients of variation (CV) of inter-day assay and intra-day assay by using Q-PCR were 1.58% and 0.88% respectively. Q-PCR could detect reproducibly 5 copies of target gene per 100 ng RNA and no pseudopositive results were found. The median NQ of PML/RARalpha mRNA was 0.450 (0.084 - 1.082) in 46 APL patients. There was no indication of any correlation of PML/RARalpha mRNA type with age, sex, hemoglobin, platelet count, percentage of promyelocytes in bone marrow detected by morphology or flow cytometry, PML/RARalpha NQ, or signs of clinically diagnosed coagulation/bleeding disorders. Compared with bcr1-form cases, bcr3-form cases had more M(3v) phenotype (42.9% vs 9.4%, P = 0.015) and higher WBC count (9.35 x 10(9)/L vs 2.15 x 10(9)/L, P = 0.038). APL cells could be classified into large side scatter population (L-SSC) and non-large side scatter population (NL-SSC) in CD45/SSC histogram of flow cytometry. 87.50% patients with bcr1-form showed L-SSC phenotype and 64.29% patients with bcr3-form showed NL-SSC phenotype. It is concluded that a sensitive Q-PCR method is established. The median NQ of PML/RARalpha mRNA was 0.450 in newly diagnosed APL patients. There was no significant difference about PML/RARalpha mRNA expression of both bcr3-form and bcr1-form APL patients. Type of PML/RARalpha transcripts is related with the morphology and immunophenotype.
Adolescent ; Adult ; Aged ; Child ; Female ; Genes, abl ; genetics ; Humans ; Leukemia, Promyelocytic, Acute ; drug therapy ; genetics ; metabolism ; Male ; Middle Aged ; Oncogene Proteins, Fusion ; analysis ; genetics ; Phenotype ; RNA, Messenger ; analysis ; genetics ; Receptors, Retinoic Acid ; analysis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; methods

Objective To establish animal models with anterior transposition of the ulnar nerve and evaluate the safety of anterior transposition of the ulnar nerve at molecular level.Methods Location of the ulnar nerve of elbow in 5 rats were found similar to human being by anatomy.Twenty healthy adult SD rats,weighting about 250 g,were performed the anterior transposition of the ulnar nerve in the right forelimbs and the left forelimbs was considered as control group.The bilateral flexor carpi ulnaris muscles were weighed and the slice of cervical spinal cord(C_6-T_1)level were prepared 1 month after the operation.Nissl staining,NADPH-d histochemical staining,IB4 staining and ChAT-immunohistochemical staining were employed to observe the spinal cord(C_6-T_1)level at molecular level;electron microscope was used to observe the ultrastructure of ChAT-positive neurons.Statistical analysis was paired T test.Results The flexor carpi ulnaris muscles in the model group(92.3±9.13mg)and control group(93.2±7.29 mg)were not significantly different(P>0.05).After anterior transposition of the ulnar nerve in rats,no significant differences in cell number and morphology in the cervical spinal cord(C_6-T_1)were found between the model group and the control group(P>0.05).No changes between the 2 groups were noted in the fine structure of anterior horn motor neurons and the expression of nenrotransmitters(P>0.05).Conclusion Anterior transposition of the ulnar nerve can be safely done in the animal models(rats).

OBJECTIVEThe aim of the study was to determine the prevalence and the distribution pattern of lesion site of intracranial vascular stenosis and to identify risk factors for the stenosis in patients with essential hypertension.

METHODSA total of 231 consecutive inpatients with essential hypertension were included in this study. Patients with the history of cerebrovascular diseases and relevant neurological symptoms were excluded. Intracranial vascular stenosis (>50% diameter reduction) was detected using CT angiography (CTA).

RESULTSOf 231 patients, 69 (29.87%) had intracranial artery stenosis. The most common stenosis site is middle cerebral artery (43.69%), followed by carotid siphon (20.39%). The stenosis in internal carotid arterial system (78.64%) was more common than in vertebrobasilar arterial system (21.56%, P < 0.05). The patients with intracranial vascular stenosis were older, had longer history of hypertension, higher levels of systolic blood pressure, higher plasma cholesterol, higher LDL-C. Lp (a), higher urinary microalbumin excretion, thicker ventricular septum, and lower levels of HDL-C than the patients without stenosis. Logistic analysis showed that systolic blood pressure (OR 1.650, 95% CI 1.134 - 2.400, P = 0.023), course of hypertension (OR 1.238, 95% CI 1.072 - 1.429, P = 0.006), LDL-C (OR 2.103, 95% CI 1.157 - 3.823, P = 0.014) and type 2 diabetes (OR 2.325, 95% CI 1.161 - 4.341, P = 0.011) were the independent risk factors of asymptomatic intracranial arterial stenosis.

CONCLUSIONSNearly 30% inpatients with essential hypertension had asymptomatic intracranial artery stenosis. The most common site of stenosis was middle cerebral artery. Hypertension, dyslipidemia and diabetes were risk factors for the development of intracranial arterial stenosis.

Aged ; Cerebral Angiography ; Female ; Humans ; Hypertension ; epidemiology ; pathology ; Intracranial Arterial Diseases ; epidemiology ; Male ; Middle Aged ; Prevalence ; Risk Factors

OBJECTIVETo explore the specificity of anti-phosphotyrosine monoclonal antibody PY20 in bcr-abl+ cells and its possible clinical applications.

METHODSBcr-abl cell lines( K562, MEG-01) and bcr-abl- cells lines( Jurkat, MCF-7 )were stained with PY20. Phosphotyrosine protein of K562 and MEG-01 cells was detected by flow cytometry before and after treatment with imatinib. Phosphotyrosine protein in bone marrow cells from 49 patients with chronic myeloid leukemia (CML), Ph+ acute lymphoblastic leukemia(Ph(+) -ALL), Ph- ALL, acute myeloid leukemia (AML-M1, M2, M3, M5, FAB classification), chronic lymphocytic leukemia (CML) and 3 normal donor. Positive cells over 5% of total cells was considered positive cases for phosphotyrosine protein. The level of tyrosine phosphorylation was determined by median fluorescence intensity (MFI).

RESULTSBcr-abl cell lines and marrow cells from 10 CML patients and 8 ALL patients were all PY20-positive, while bcr-abl- cell lines and marrow cells from 18 leukemia patients and 3 normal donor were all PY20-negative. MFI decreased remarkably after blocked by imatinib in K562 cells and MEG-01 cells. The positive cell percent of marrow cells from 10 newly diagnosed CML patients and 9 imatinib-sensitive CML patients was (54.20 +/- 19.82)% and (14.84 +/- 6.17)% (P < 0.05), while that of 2 cases of imatinib-resistant was 64.3% and 57.2%. There was significant difference of MFI between imatinib-resistant patients and imatinib-sensitive patients (99.42 +/- 4.87 vs 46.41 +/- 4.67, P < 0.01).

CONCLUSIONPY20 monoclonal antibody is highly specific for bcr-abl+ cells. It might be useful in rapid detection of bcr-abl+ cells and sensitivity to imatinib of CML patients.

Antibodies, Monoclonal ; analysis ; Bone Marrow Cells ; metabolism ; Flow Cytometry ; Fusion Proteins, bcr-abl ; analysis ; Humans ; K562 Cells ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; metabolism ; Leukemia, Myeloid, Acute ; metabolism ; Phosphotyrosine ; analysis ; immunology

BACKGROUNDReal-time quantitative RT-PCR (RQ-PCR) assay has become a vital tool to monitor residual disease of leukemia. However, the complexity and standardization of RQ-PCR should never be overlooked and the results should be interpreted cautiously in clinical conditions. We aimed to assess the methodology of RQ-PCR and its clinical applications in monitoring molecular kinetics of 36 newly diagnosed cases of acute promyelocytic leukemia patients with t (15; 17) from October 2004 to December 2005.

METHODSAll the TaqMan probe-based RQ-PCR reactions and analysis were performed on an ABI-PRISM 7,500 platform. The quantitation of PML-RARalpha transcripts was represented by the normalized quotient, that is, PML-RARalpha transcript copies divided by ABL transcript copies. According to induction therapy, the patients were classed into two groups: group 1 (n = 23), three-drug combination including arsenics, all-trans retinoic acid and mitoxantrone; and group 2 (n = 13), two-drug combination from all-trans retinoic acid, arsenics and mitoxantrone.

RESULTSThe sensitivity of RQ-PCR was 1 per 10(5) cells and 5 copies of the PML-RARalpha transcript could be reproducibly detected. No false positive results occurred in 40 non-acute promyelocytic leukemia samples. Optimal amplification efficiency could be attained, which was determined by the slope of the standard curves (slope: -3.2 - -3.7). The inter-assay and intra-assay variation coefficients of the method were 1.01% and 0.56% respectively. Although the time to attain hematological complete remission was similar in both groups, the time to achieve molecular remission of group 1 was significantly shorter than that of group 2 (61 days vs 75 days, P = 0.034). The rate of molecular remission within 70 days was higher in group 1 than in group 2 (75.00% vs 38.46%, P = 0.036). Compared with pretreatment, median reduction of the PML-RARalpha transcript before first consolidation therapy differed significantly between group 1 and group 2 (log scale, 3.15 vs 2.31, P = 0.024). Interestingly, we found that PML-RARalpha transcript levels temporarily increased in bone marrow (7 patients) and peripheral blood (22 patients) samples of patients during induction therapy in both groups.

CONCLUSIONSThe RQ-PCR assay is reliable for the detection of PML-RARalpha transcripts. Arsenics, all-trans retinoic acid and mitoxantrone triad induction treatment of acute promyelocytic leukemia is superior to two-drug combination induction therapy in terms of the molecular response.

Adolescent ; Adult ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Child ; Female ; Humans ; Leukemia, Promyelocytic, Acute ; blood ; drug therapy ; genetics ; Male ; Middle Aged ; Oncogene Proteins, Fusion ; genetics ; RNA, Messenger ; analysis ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Sensitivity and Specificity

Objective Epidemiological studies have suggested that alcohol drinking is closely associated with di-verse tumor development, but fewer laboratory studies or mechanism analysis is made. Our study aims to explore the effect of alcohol on the tumorigenicity and metastasis of breast cancer in nude mice in vivo and on the malignant behavior in vitro, and whether it is related to endoplasmic reticulum stress ( ERS) . Methods We first established the nude mouse model of breast cancer with chronic alcohol consumption (2%) and to detect the effects of ethanol on tumor growth and metastasis. We also observed the changes of malignant biological behavior of EO771 cells in cell culture induced by chronic alcohol treatment (0. 2%) using MTT, Transwell and soft agar tests, and detect the expression of ROS and ERS marker, i. e. p?eIF2a, Bip, and XBP1?s. Results The in vivo experiment showed that alcohol drinking enhanced the growth and metasta?ses of breast cancer in nude mice. The in vitro test showed that chronic exposure to ethanol also promoted the cell prolifera?tion and migration ability, increased the ROS expression, activated ERS pathway, and enhanced the expression of p?eIF2a, Bip and XBP1?s. Conclusions Alcohol drinking may promote the malignant behavior of breast cancer cells through endo?plasmic reticulum stress.

OBJECTIVETo summarize the experience of diagnosis and treatment in solid-pseudopapillary tumor of pancreas (SPT).

METHODRetrospectively analyzed 12 cases of SPT diagnosed between 2000 August and 2004 February in Peking Union Medical College Hospital.

RESULTSThe average age of these 12 cases was 29.1, including 11 women and 1 men. The tumors of 6 cases were located in the pancreatic head, one tumor was in the neck of pancreas and 5 tumors were in the body and tail of pancreas. There were no characteristic clinical manifestations. Solid and solid-cystic masses of low echo were found in pancreas in ultrasonic examinations. CT scan found masses of low density in pancreas, while irregular enhancement appeared in the circumference of all tumors in enhanced CT scan sequences. Tumor markers in patients' serum were all negative. Pancreaticoduodenectomy were performed in 2 cases. Resection of tumors and pancreaticojejunostomy were performed in 3 cases. Simple resection of tumor was performed in 1 case. Segmental pancreatectomy and pancreaticojejunostomy was performed in one patient. Five patients underwent distal pancreatectomy and spleen resection. Ten patients were followed up with the average time of 14.3 months. No evidence of relapses and metastasis in these cases was found.

CONCLUSIONSolid-pseudopapillary tumor of pancreas was one of rare pancreatic neoplasm with low malignant potential, affecting primarily young women. Complete resection resulted in long-term survival.

Adolescent ; Adult ; Carcinoma, Papillary ; diagnosis ; therapy ; Female ; Humans ; Male ; Middle Aged ; Pancreatectomy ; Pancreatic Neoplasms ; diagnosis ; therapy ; Pancreaticoduodenectomy ; Retrospective Studies ; Splenectomy ; Tomography, X-Ray Computed