Multi-color super-resolution imaging to study human coronavirus RNA during cellular infection

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the third human coronavirus within 20 years that gave rise to a life-threatening disease and the first to reach pandemic spread. To make therapeutic headway against current and future coronaviruses, the biology of coronavirus RNA during infection must be precisely understood. Here, we present a robust and generalizable framework combining high-throughput confocal and super-resolution microscopy imaging to study coronavirus infection at the nanoscale. Employing the model human coronavirus HCoV-229E, we specifically labeled coronavirus genomic RNA (gRNA) and double-stranded RNA (dsRNA) via multicolor RNA-immunoFISH and visualized their localization patterns within the cell. The exquisite resolution of our approach uncovers a striking spatial organization of gRNA and dsRNA into three distinct structures and enables quantitative characterization of the status of the infection after antiviral drug treatment. Our approach provides a comprehensive framework that supports investigations of coronavirus fundamental biology and therapeutic effects.

Impacts of A20 gene deletion on clinicopathological features and prognosis of diffuse large B cell lymphoma and relative molecular mechanism / 重庆医学

Objective To detect the A20 gene deletion,investigate the impacts of A20 gene deletion on clinicopathological features and prognosis of DLBCL,and relationship between activation of NF-κB pathway and relative molecular pathogenesis.Methods A20 gene deletion was detected by fluorescence in situ hybridization (FISH).The expression of A20,Survivin,P65 and Ki-67 were detected by immunohistochemistry stain.Apoptosis was assayed by TUNEL.Follow-up and statistical analysis were done.Results The deletion rate of A20 gene was 21.7％.The deletion rate of A20 gene was obviously higher in ABC-like DLBCL than that in GCB-like DLBCL (30.6％ vs.8.3％,P＜0.05).It was observed that there was a negative correlation between A20 protein expression and A20 gene deletion (r=-0.259,P =0.023).The expression of P65 and Survivin protein was positively correlated with the A20 gene deletion (r=0.280,P =0.015;r =0.313,P =0.007).Apoptosis rate was significantly reduced in DLBCL patients with A20 gene deletion.The apoptosis rate was higher in cases with positive expression of A20 protein,while that was lower in cases with positive expression of p65 and Survivin protein than those with negative expression of corresponding protein.There was no statistically significant difference in apoptosis rate between ABC-like and GCB-like DLBCL patients (P＞0.05).COX regression analysis indicated that age,A20 gene deletion,types of DLBCL and Ki67 expression were independent factors associated with survival status.Log-rank test showed that there was a statistical difference in survival status between the cases with and without A20 gene deletion (P=0.015).Conclusion A20 gene deletion may associate with the attenuation of A20 protein expression.The latter weakens negative feedback regulation of A20 protein for NF-κB pathway.An up-regulated expression of Survivin and abnormal proliferation and apoptosis may be result from the abnormal activation of NF-κB.A20 gene deletion brings certain influence on clinical course and prognosis of DLBCL.

Vincristine reduces A20 siRNA-induced proliferation in diffuse large B cell / 基础医学与临床

Objective To investigate the impact of MDR1-targeting small interfering RNA (siRNA) on diffuse large B-cell lymphoma cell OCI-LY1 proliferation.Methods A20 gene was silenced using RNA interference.An optimal concentration and treatment duration of vincristine were selected using MTT.Before and after siRNA transfection, proliferation of OCI-LY1 cells was assayed using MTT assay, and cellular apoptosis was detected using FCM before or after the treatment of the cells with VCR.Detection of A20, NF-κB (p65) and Pgp proteins were conducted using Western blot whereas mRNA of the A20 and MDR1 genes were examined using real time PCR.Results1)Proliferation of OCI-LY1 cells was enhanced (P<0.001) after the transfection with siRNA-2,(P<0.05).In addition, cell proliferation curve was declined after VCR stimulation, but the decrease was slower in siRNA-transfected cells than the untransfected counterparts.2)Apoptostic rate was lower in siRNA-transfected cells than theuntransfected counterparts, and the rate was higher in the cells after treatment with the drug for 24 h (P<0.05).Increased apoptosis was more obvious in control OCI-LY1 cells than in siRNA-transfected cells after treatment with VCR(P<0.05).3)The expression of MDR1 mRNA and Pgp (P<0.001) was significantly increased after transfection, but the expression of MDR1 mRNA and Pgp were significantly decreased (P<0.05).The expression in VCR group was significantly lower than that in siRNA-transfected cells+VCR group (P<0.01).ConclusionsA20 siRNA could effectively enhance NF-kappa B expression in OCI-LY1 cells.NF-kappa B may up regulate the expression of its downstream genes such as MDR1 and cause apoptosis, in turn enhancing the inhibition of cell proliferation.VCR can reduce MDR1 mRNA and Pgp expression in OCI-LY1 cells and the effect of VCR could be attenuated by A20 siRNA.

Characteristics and impact factors of ocular damage effects induced by transitional near-infrared laser:a review / 军事医学

The transitional near-infrared (NIR) laser was defined as ranging from 1.3μm to 1.4μm, within which the most sensitive tissue to laser damage changed from the retina to the cornea.The ocular damage effect has attracted much attention due to the increased varieties and output power of laser in this spectrum region in recent years.Compared with visible and mid-and-far infrared wavelengths, the ocular damage effect induced by transitional NIR wavelengths has many peculiarities and impact factors due to the bulk absorptionby ocular media.This paper reviews the existing ocular damage threshold data and analyzes the characteristics, impact factors and unresolved issues relating to ocular effects induced by laser radiation over the transition zone.

The role of GRP78 on the resistance to cisplatin in SPCA-1 cell line / 中国肺癌杂志

<p><b>BACKGROUND AND OBJECTIVE</b>The expression levels of glucose-regulated protein 78 (GRP78) were elevated and correlated with resistance to chemotherapy drug VP-16 in lung cancer cells. However, little is known about the relationship between its expression and resistance to cisplatin in lung cancer cells. The aim of this study was to investigate the expression of GRP78 under the induction of A23187 and its significance in the resistance to anti-tumor drugs cisplatin in a human lung cancer SPCA-1 cell line.</p><p><b>METHODS</b>RT-PCR and Western blot were used to analyze the expression of GRP78 at mRNA and protein levels in SPCA-1 cell line induced byA23187 at different concentrations (0, 1, 2, 4, 6 microM). MTT was used to determine the effect of cisplation on cell survival.</p><p><b>RESULTS</b>The expressions of GRP78 at both mRNA and protein levels were increased obviously in SPCA-1 cell line induced by A23187, with a manner of dose-dependent of A23187 to a great degree; MTT assay showed that the cell survival rate of the A23187-induced group decreased significantly compare to the control group, also with a concentration-dependent manner of A23187.</p><p><b>CONCLUSION</b>The expression of GRP78 at both mRNA and protein levels were increased obviously in SPCA-1 cell line induced by A23187. The enhancement of GRP78 showed a negative correlation with the cell survival rate treated by cisplatin. All these indicated that overexpression of GRP78 can enhance the sensitivity to cisplatin and there is correlation between the expression of GRP78 and resistance to cisplatin of human lung cancer SPCA-1 cell line.</p>

Application of microfluidic gradient chip in the analysis of lung cancer chemotherapy resistance.

The major challenge of chemotherapy is the disease resistance for many lung cancer patients. Integrated microfluidic systems offer many desirable characteristics and can be used in cellular biological analysis. This work aimed to study the correlation between the expression of Glucose Regulated Protein-78 (GRP78) and the resistance to anticancer drug VP-16 in human lung squamous carcinoma cell line SK-MES-1 using an integrated microfluidic gradient chip device. We used A23187, a GRP78 inducer, with a gradient concentration in the upstream network of the device to induce the expression of GRP78 in the cells cultured in the downstream before the addition of VP-16. The expression of GRP78 was detected by immunofluorescence, the apoptosis for the cells treated by VP-16 was assessed morphologically by 4',6-diamidino-2-phenylindole (DAPI) staining. The results indicated that the expressions of GRP78 increased greatly for the cells under the induction of A23187 with a dose-depended manner, while the percentage of apoptotic cells decreased significantly after being treated by VP-16. Our results from this study confirmed the role of GRP78 played in the chemotherapy resistance to VP-16 in SK-MES-1 cell line, suggesting that the integrated microfluidic systems may be an unique approach for characterizing the cellular responses.

Relationship between coronary atherosclerotic stenosis and cerebral atherosclerotic stenosis / 老年心脏病学杂志（英文版）

To investigate the relationship between severity of cerebrovascular atherosclerosis stenosis and that of coronary atherosclerosis stenosis.Methods Cerebral angiography and coronary angiography were performed in 34 patients who had coronary disease with cerebral ischemia.Patients were divided into 3 subgroups according to the degree ofstenosis on angiography,concomitant diseases,risk factors and biochemical data.Results The follow-up study showed that the incidence of cardiac and cerebrovascular death increased significantly in patients with moderate to severe stenosis of coronary and cerebral arteries;the severity of stenosis in the coronary artery parallels that in the solitary carotid artery,or dual carotid and vertebral arteries.Conclusions Patients with coronary and cerebral artery stenosis,especially those with multi-risk factors,such as hypertension,diabetes and cigarette smoking,should receive intensive treatment to reduce cardiac and cerebrovascular events.(J Geriatr Cardiol 2008;5:227-229)

Comparison of multi-slice CT coronary artery imaging with coronary angiography / 中国组织工程研究

BACKGROUND: Primary studies suggest that coronary artery stenosis is highly exactly shown by 16-slice spiral CT coronary artery imaging.OBJECTIVE: To compare the accuracy and limitation between coronary angiography and multi-slice computed tomography (MSCT) coronary artery imaging to diagnose moderate and severe coronary artery stenosis. DESIGN, TIME AND SETTING: Clinical diagnostic study based on gold standard, which was carried out in the Department of Cardiology, Xuanwu Hospital, Capital Medical University from June 2005 to March 2006. PARTICIPANTS: Twenty-eight patients with suspected coronary arteriosclerotic heart disease were examined by 64-slice spiral CT coronary artery imaging and coronary artery angiography during the 1-month hospitalization in the Department of Cardiology, Xuanwu Hospital, Capital Medical University from June 2005 to March 2006. METHODS: 280 segments of 28 patients were quantitatively analyzed coronary artery stenosis by selective coronary artery angiography and multi-slice spiral CT imaging based on eye-measurement diameter method. MAIN OUTCOME MEASURES: True positive, true negative, false positive, false negative, sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of coronary artery stenosis were measured by multi-slice spiral CT imaging.RESULTS: All 28 patients were included in the final analysis. Multi-slice spiral CT imaging showed that the sensitivity, specificity, positive predictive value, and negative predictive value were 46.5%, 97.6%, 86.8%, and 84.3%, respectively. If excluding the effect of 31 coronary segments with severe calcification, the sensitivity, specificity, positive predictive value and negative predictive value were 66.7%, 98.6%, 90.3% and 93.6%, respectively.CONCLUSION: Multi-slice spiral CT imaging is simple, reliable, noninvasive and safe; moreover, it has a good potential for diagnosing especially excluding coronary arteriosclerotic heart disease, but still some limits.