Influence of Osteopontin Short Hairpin RNA on the Proliferation and Activity of Rat Vascular Smooth Muscle Cells

To investigate the influence of osteopontin （OPN） short hairpin RNA （shRNA） on the proliferation and activity of rat vascular smooth muscle cells （VSMCs）, the expressing vector of shRNA targeting OPN was constructed and transferred into the rat VSMCs. After amplification and purification, pGenesil-1/OPNshRNA1 （PG1）, pGenesil-1/OPNshRNA2 （PG2） and pGenesil-1/OPNshRNAHK （PGH） were transfected into the cultured rat VSMC by LipofectamineTM 2000. Transfected cells were visualized by using an inverted fluorescent microscope. VSMCs transfected by optimal recombined plasmid was selected by culturing in G418 48 h later. Nude cells and cells transfected by PGH were used as control. The expression levels of OPN mRNA and protein were assayed by RT-PCR and Western blotting. The OPN of VSMCs was suppressed by transfection of optimal recombined plasmid, and the changes in cell proliferation, adhesion and motility were evaluated by MTT, adhesion test and transwell chamber test. Levels of type I and Ⅲ collagen were measured with ELISA kit. Our results showed that VSMCs stably transfected by OPN shRNA accounted for over 50% of total cells. OPN mRNA and protein were reduced by 81% and 67% （P〈0.01） by PG1, 73% and 52% （P〈0.01） by PG2, respectively while no change was found in PGH and non-treated VSMCs. PG1 significantly suppressed the proliferation, adhesion, mobility of VSMCs and reduced the amount of type Ⅰ and Ⅲ collagen. It is concluded that recombinant plasmid can be success-fully transfected into VSMCs by LipofectamineTM 2000 and inhibit the expression of OPN. The proliferation, adhesion and mobility of VSMCs can be inhibited by knocking down OPN expression. Moreover, the transferring capability of cells is attenuated, and the secretion of type Ⅰ and Ⅲ collagen is inhibited aftter knocking-down of OPN expression. The study provides experimental evidence for clinical prevention of restenosis after percutaneous coronary intervention （PCI） by RNA interference （RNAi） technology.

Clinicopathological Analysis of Central and Extraventricular Neurocytoma:A Report of 17 Cases

Neurocytoma,a rare brain tumor,is characterized by a mass located mainly in cerebral ventricles.It is prone to be misdiagnosed as oligodendroglioma or ependymoma due to their similar histopathological features in clinical practice.This study aimed to examine the clinicopathological features and differential diagnosis of central and extraventricular neurocytoma.The clinical and histopathological data of 17 patients (male:female=7:10;age:4-41 years;mean age:27.4 years) with central or extraventricular neurocytoma were retrospectively analyzed.These patients showed typical radiological,histopathological and immunohistochemical features of neurocytoma.The tumor tissue was found to be composed of small uniform cells with round nuclei and clear cytoplasm resembling that of oligodendroglioma and ependymoma.Immunohistochemistry revealed the tumor tissues were positive for neuronal markers such as synaptophysin (SYN) and neuronal nuclear antigen (NeuN).It was concluded histopathological features of neurocytoma overlaps with some tumors in the central neural system.Immunopositivity for SYN and NeuN can help differentially diagnose neurocytoma.

Effects of Aconitine on [Ca^(2+)] Oscillation in Cultured Myocytes of Neonatal Rats

In order to investigate the effects of aconitine on [Ca2＋] oscillation patterns in cultured myocytes of neonatal rats, fluorescent Ca2＋ indicator Fluo-4 NW and laser scanning confocal micro- scope （LSCM） were used to detect the real-time changes of [Ca2＋] oscillation patterns in the cultured myocytes before and after aconitine （1.0 μmol/L） incubation or antiarrhythmic peptide （AAP） and aconitine co-incubation. The results showed under control conditions, [Ca2＋] oscillations were irregu- lar but relatively stable, occasionally accompanied by small calcium sparks. After incubation of the cultures with aconitine, high frequency [Ca2＋] oscillations emerged in both nuclear and cytoplasmic regions, whereas typical calcium sparks disappeared and the average [Ca2＋] in the cytoplasm of the cardiomyocyte did not change significantly. In AAP-treated cultures, intracellular [Ca2＋] oscillation also changed, with periodic frequency, increased amplitudes and prolonged duration of calcium sparks. These patterns were not altered significantly by subsequent aconitine incubation. The basal value of [Ca2＋] in nuclear region was higher than that in the cytoplasmic region. In the presence or absence of drugs, the [Ca2＋] oscillated synchronously in both the nuclear and cytoplasmic regions of the same cardiomyocyte. It was concluded that although oscillating strenuously at high frequency, the average [Ca2＋] in the cytoplasm of cardiomyocyte did not change significantly after aconitine incuba- tion, compared to the controls. The observations indicate that aconitine induces the changes in [Ca2＋] oscillation frequency other than the Ca2＋ overload.

Essential oil from Saussurea costus inhibits proliferation and migration of Eca109 cells via mitochondrial apoptosis and STAT3 signaling

Objective:To investigate the effect and its underlying molecular mechanisms of essential oil from Saussurea costus in esophageal cancer cell line Eca109.Methods:The chemical composition of essential oil from Saussurea costus was investigated by gas chromatography-mass spectrometry(GC-MS).The anti-proliferative,anti-migrative,and apoptotic effects of essential oil from Saussurea costus against Eca109 cells were analyzed.Moreover,the expression of proteins associated with cell cycle,metastasis,and apoptosis was determined.Results:GC-MS analysis showed that essential oil from Saussurea costus was predominantly comprised of sesquiterpenes.Saussurea costus essential oil inhibited the viability of Eca109 cells in a dose-and time-dependent manner with IC_(50) values of(24.29±1.49),(19.16±2.27)and(6.97±0.86)μg/mL at 12,24,and 48 h,respectively.The expression levels of target proteins in the cell cycle(phase G_(1)/S),including cyclin D1,p21,and p53,were affected by Saussurea costus essential oil.The essential oil also downregulated the expression of metastasis-related proteins MMP-9 and MMP-2.Moreover,it induced apoptosis of Eca109 cells through the mitochondrial pathway,as well as inhibition of STAT3 phosphorylation.Conclusions:The essential oil from Saussurea costus exhibited anti-proliferative,anti-migrative,and apoptotic effects on Eca109 cells,and could be further explored as a potential anti-esophageal cancer agent.

Clinicopathological and immunohistochemical features of pilomyxoid astrocytoma: a report of six cases

Objective： The aim of this study was to study the clinicopathological and immunohistochemical features of pilo- myxoid astrocytoma （PMA）. Methods： The clinical and pathologic features in six cases of PMA were analyzed. Immunohisto- chemical staining for glial fibrillary acidic protein （GFAP）, synaptophysin （Syn）, Chromogranin A （CgA）, cytokeratin （AEI/AE3）, epithelial membrane antigen （EMA） and Ki67 was performed on paraffin-embedded sections. Results： Among the six cases, five occurred in female patients, one was male, the age at diagnosis ranged from 2 to 15 years. Four cases were located in the hypothalamic area and optic pathway, one case in the third ventricle, and one case in left parietal lobe. On imaging, PMAs often appears as well-circumscribed mass. Microscopically, the tumor was composed of monomorphous bipolar （piloid） cells setting in a prominent myxoid background with an angiocentric radiating growth pattern in some areas. PMA lacked biphasic pattern, Rosenthal fibers and eosinophilic granular bodies which were usually typical in a classic pilocytic astrocytoma （PA）. Immunohistochemcal study showed that the tumor cells were diffusely positive for GFAP. Syn positive staining was observed in one case. The Ki67 labeling index measured less than 5%. Conclusion： PMA is a distinct aggressive variant of pilocytic astrocytoma with special histological and immunohistochemical features. It is typically a rare tumor of early childhood. Im- munohistochemical staining for GFAP and Syn is helpful in differential diagnosis.

Effects of Dual-Perspective Nursing Intervention Based on Safety and Comfort Needs on Psychological Status and Safety Quality of MRI Patients

Objective:To investigate the effect of dualperspective nursing intervention based on safety and comfort needs on the psychological status and safety quality of patients undergoing magnetic resonance imaging(MRI).Methods:A total of 100 patients who received MRI in our hospital from October 2017 to December 2019 were selected and randomly divided into two groups,50 cases for each group.Routine nursing intervention was performed in the control group,and double-perspective nursing intervention based on safety and comfort needs in addition to routine nursing was performed in the observation group.Psychological status and examination of safety quality were observed and compared between the two groups.Results:The scores of self-rating anxiety scale(SAS)and self-rating depression scale(SDS)in the observation group were lower than those in the control group after intervention.Meanwhile,the scores of early treatment of dangerous conditions,implementation of safety measures,prevention of accidental injuries,communication of safety information,education of safety knowledge,standardized examination and awareness of self-safety management in the observation group were higher than those in the control group after intervention,and the differences were statistically significant(P<0.05).Conclusion:Dual-perspective nursing intervention based on safety and comfort needs is effective for patients undergoing MRI examination,which can improve the psychological status of patients and improve the quality of examination safety.

Analysis of the Application Value of MRI and CT Diagnosis of Lumbar Disc Herniation

Objective:To analyze the application value of MRI and CT diagnosis for those with lumbar disc herniation.Methods:The data of 62 patients with LDH treated in our hospital from January 2019 to December 2019 were analyzed retrospectively.All patients were diagnosed as LDH after operation,and 62 patients were treated with MRI and CT.The accuracy of CT and MRI in the diagnosis was analyzed according to the gold standard of operation results.Results:62 patients were diagnosed as LDH after operation,including 25 cases with intervertebral disc herniation,12 cases with intervertebral disc prolapse,8 cases with intervertebral disc nodule,7 cases with intervertebral disc dissociation as well as 10 cases with intervertebral disc bulge.The accuracy of MRI diagnosis(95.16%)was higher than that of CT(75.81%),and the difference was statistically significant(P<0.05).Conclusion:For the diagnosis of LDH,MRI is more accurate,but CT is faster and cheaper.Both of them have their own advantages can be chosen clinically according to the actual situation of patients on the basis of ensuring the diagnostic accuracy.

The Efficacy of Different Interventiona|Che moembolization Regiments Combined with Radiofrequency Ablation in the Treatment of Primary Liver Cancer and its Effects on Patients’Immune Function

Objective:To study and compare the clinical effects of the combination of different interventional embolization chemotherapy and radiofrequency ablation in patients with primary liver cancer.Methods:In this paper,based on the target data validation of those 60 patients with primary liver cancer treated in the hospital during the period from May 2017 to May 2018,the double-blind method was used for the comparison between groups.Patients in the reference group were treated with the combination of cisplatinum interventional chemoembolization regimen and radiofrequency ablation,while those in the experimental group were treated with the combination of lobaplatin interventional chemoembolization regimen and radiofrequency ablation.Then,the efficacy of both groups was compared.Results:After the treatment,the clinically effective total value,the calculation value of adverse reactions,and the value of IgA,IgM,IgG and AFP of the experimental group were compared with those of the reference group.In addition,the value of IgA,IgM,IgG and AFP of both groups after and before the treatment were compared.The experimental results showed that the data was statistically significant(P<0.05).Conclusion:The combination of lobaplatin and cisplatin interventional embolization chemotherapy and radiofrequency ablation has some effects in patients with primary liver cancer,but the former one showed more significant effects.

Comparison of the Values of MRI in the Differential Diagnosis of Symptomatic Carotid Stenosis and Atherosclerotic Plaque

Objective:To evaluate the values of Carotid Magnetic Resonance Imaging(MRI)in the differential diagnosis of symptomatic carotid stenosis and atherosclerotic plaque.Methods:56 patients with ischemic cerebrovascular disease admitted in our hospital from October 2018 to October 2019 were selected and treated with Carotid MRI and digital subtraction angiography(DSA)examinations.According to the two examination results recorded and the“gold standard”of DSA examination,values of Carotid MRI in the differential diagnosis of symptomatic carotid stenosis were evaluated.Results:According to the“gold standard”of DSA examination,the sensitivity and specificity of MRI examination for carotid stenosis were:Mild:92.54%and 97.78%;Moderate:85.71%and 88.78%;Severe:100.00%and 97.8%;and complete occlusion:100.00%and 100.00%;The proportions of intraplaque haemorrhage and ruptured fibrous cap in different degrees of carotid artery stenosis were:Mild:30.16%and 22.22%;Moderate:43.48%and 39.13%;And severe:57.89%and 52.63%.Conclusion:MRI examination can evaluate the degree of symptomatic carotid artery stenosis,and show the characteristics of atherosclerotic plaque at the same time to provide a reference for early clinical differential diagnosis and treatment.

The precursor frequency of alloreactive CTLs is proportional to the number of T cell epitope specificities

The aim of this study is to find the experimental evidence that the precursor frequency of alloreactive CTLs is proportional to the number of the T-cell epitope specificities. The number of T-cell epitope specificities was manipulated by pulsing different humor of HLA-A2 restricted peptide（s） onto the T2 cells, which acted as stimulating cells to elicit allo-reaction by co-culturing with peripheral blood lymphocytes （PBLs） of HLA-A2 negative individual. Ten HLA-A2 restricted peptides （all were normal cell components ） were synthesized, and cell peptide extract was prepared by frozen and thawed. T2 cells loaded with different number of peptide（s） were co-cultured with PBLs of an HLA-A2 negative individual; the latter were stained with PKH67 in advance. Then the proliferation was monitored with flow cytometry, and the precursor frequency of the effector cells was ＇analyzed by the ModFit Software. After 6 d of culture, no proliferation was observed in the bulk culture of PBL alone, and obvious proliferation took place when PBLs of the HLA-A2 negative were co-cultured with T2 cells loaded with or without loading peptide（ s）. The precursor frequency of the alloreactive CTLs was 0.052 819 for co-culture with T2 cells loaded without peptide; however it was 0. 030 429 for T2 cells with EBV/LMP2A and 0. 030 528 for T2 cells loaded with a single autogeneic peptide, and increased up to 0. 144 942 for T2 cells loaded with 10 autogeneic peptides; the precursor frequency was 0. 203 649 when co-cultured with T2 cells loaded with miscellaneous peptides extracted from the cytoplasm of T2 cells. This study reveals that the precursor frequency of alloreactive CTLs is proportional to the number of T-cell epitope specificities, and independent of the density of the allogeneic HLA Class Ⅰ molecule. Our findings support the hypothesis that the alloreactive T cell populations comprise miscellaneous T cell clones; each is specific to corresponding pMHC. The novel constellation of peptides presented by allogeneic MHC molecules makes thousands of different epitopes, which account for the exceptional high precursor frequency of alloreactive T cells.

Treatment Strategies for Huge Central Neurocytomas

Central neurocytomas（CNs）, initially asymptomatic, sometimes become huge before detection. We described and analyzed the clinical, radiological, operational and outcome data of 13 cases of huge intraventricular CNs, and discussed the treatment strategies in this study. All huge CNs（n=13） in our study were located in bilateral lateral ventricle with diameter ≥5.0 cm and had a broad-based attachment to at least one side of the ventricle wall. All patients received craniotomy to remove the tumor through transcallosal or transcortical approach and CNs were of typical histologic and immunohistochemical features. Adjuvant therapies including conventional radiation therapy（RT） or gamma knife radiosurgery（GKRS） were also performed postoperatively. Transcallosal and transcortical approaches were used in 8 and 5 patients, respectively. Two patients died within one month after operation and 3 patients with gross total resection（GTR） were additionally given a decompressive craniectomy（DC） and/or ventriculoperitoneal shunt（VPS） as the salvage therapy. Six patients received GTR（＋RT） and 7 patients received subtotal resection（STR）（＋GKRS）. Eight patients suffered serious complications such as hydrocephalus, paralysis and seizure after operation, and patients who underwent GTR showed worse functional outcome [less Karnofsky performance scale（KPS） scores] than those having STR（＋GKRS） during the follow-up period. The clinical outcome of huge CNs seemed not to be favorable as that described in previous reports. Surgical resection for huge CNs should be meticulously considered to guarantee the maximum safety. Better results were achieved in STR（＋GKRS） compared with GTR（＋RT） for huge CNs, suggesting that STR（＋GKRS） may be a better treatment choice. The recurrent or residual tumor can be treated with GKRS effectively.