The mechanical properties of cell nuclei have been recognized to reflect and modulate important cell behaviors such as migration and cancer cell malignant tendency. However, these nuclear properties ...are difficult to characterize accurately using conventional measurement methods, which are often based on probing or deforming local sites over a nuclear region. The corresponding results are sensitive to the measurement position, and they are not decoupled from the cytoplasmic properties. Microfluidics is widely recognized as a promising technique for bioassay and phenotyping. In this report, we develop a simple and nondestructive approach for the single-cell quantification of nuclear elasticity based on microfluidics by considering different deformation levels of a live cell captured along a confining microchannel. We apply two inlet pressure levels to drive the flow of human nasopharyngeal epithelial cells (NP460) and human nasopharyngeal cancerous cells (NPC43) into the microchannels. A model considering the essential intracellular components (cytoplasm and nucleus) for describing the mechanics of a cell deforming along the confining microchannel is used to back-calculate the cytoplasmic and nuclear properties. On the other hand, we also apply a widely used chemical nucleus extraction technique to examine its possible effects (
e.g.
, reduced nuclear modulus and reduced lamin A/C expression). To determine if the decoupled nuclear properties are representative of cancer-related attributes, we classify the NP460 and NPC43 cells using the decoupled physical properties as classification factors, resulting in an accuracy of 79.1% and a cell-type specificity exceeding 74%. It should be mentioned that the cells can be recollected at the device outlet after the nondestructive measurement. Hence, the reported cell elasticity measurement can be combined with downstream genetic and biochemical assays for general cell research and cancer diagnostic applications.
Nondestructive quantification of cytoplasm and nucleus elasticity based on multiple levels of cell deformation.
Abstract EBV-associated human malignancies may originate from B cells and epithelial cells. EBV readily infects B cells in vitro and transforms them into proliferative lymphoblastoid cell lines. In ...contrast, infection of human epithelial cells in vitro with EBV has been difficult to achieve. The lack of experimental human epithelial cell systems for EBV infection has hampered the understanding of biology of EBV infection in epithelial cells. The recent success to infect human epithelial cells with EBV in vitro has allowed systematic investigations into routes of EBV entry, regulation of latent and lytic EBV infection, and persistence of EBV infection in infected epithelial cells. Understanding the biology of EBV infection in human epithelial cells will provide important insights to the role of EBV infection in the pathogenesis of EBV-associated epithelial malignancies including nasopharyngeal carcinoma and gastric carcinoma.
Aims This study aimed at evaluating the potential anti-proliferative effects of the microRNA let-7 family in nasopharyngeal carcinoma (NPC) cells. In addition, the association between let-7 ...suppression and DNA hypermethylation is examined. Materials and methods Levels of mature let-7 family members (-a, -b, -d, -e, -g, and -i) in normal nasopharyngeal cells (NP69 and NP460) and nasopharyngeal carcinoma cells (HK1 and HONE1) were measured by real-time quantitative PCR. Cell-proliferation assay and c-Myc immunohistochemical staining were performed on NPC cells transfected with let-7 precursor molecules. In addition, expression changes in let-7 family members in response to demethylating agents (5-azacytidine and zebularine) were also examined. Results In comparison with the normal nasopharyngeal cells, let-7 (-a, -b, -d, -e, -g, and -i) levels were reduced in nasopharyngeal carcinoma cells. Ectopic expression of the let-7 family in nasopharyngeal carcinoma cells resulted in inhibition of cell proliferation through downregulation of c-Myc expression. Demethylation treatment of nasopharyngeal carcinoma cells caused activation of let-7 expression in poorly differentiated nasopharyngeal carcinoma cells only. Conclusion Our results suggested that miRNA let-7 might play a role in the proliferation of NPC. DNA methylation is a potential regulatory pathway, which is affected when let-7 is suppressed in NPC cells. However, the extent of DNA hypermethylation/hypomethylation in regulating let-7 expression requires further elucidation.
Background and Aims:
Data on the natural history of elderly-onset ulcerative colitis UC are limited. We aimed to investigate clinical features and outcomes of patients with elderly-onset UC.
Methods:
...Patients with a confirmed diagnosis of UC between 1981 and 2013, from 13 hospitals within a territory-wide Hong Kong Inflammatory Bowel Disease Registry, were included. Clinical features and outcomes of elderly-onset patients, defined as age ≥ 60 years at diagnosis, were compared with those of non-elderly-onset disease < 60 years at diagnosis.
Results:
We identified 1225 patients, of whom 12.8% 157/1225; 56.1% male had elderly-onset UC. Median duration of follow-up was 11 years interquartile range, 6–16 years. Age-specific incidence of elderly-onset UC increased from 0.1 per 100000 persons before 1991 to 1.3 per 100000 persons after 2010. There were more ex-smokers 32.2% vs. 12.2%, p < 0.001 and higher proportion of comorbidities p < 0.001 in elderly-onset than non-elderly-onset patients. Disease extent, corticosteroids, immunosuppressants use, and colectomy rates were similar between the two groups. Elderly-onset disease was an independent risk factor for cytomegalovirus infection odds ratio 2.9, 95% confidence interval 1.6–5.2, p < 0.001. More elderly-onset patients had Clostridium difficile infection 11.0% vs. 5.4%, p = 0.007, hospitalisation for UC exacerbation 50.6% vs. 41.8%, p = 0.037, colorectal cancer 3.2% vs. 0.9%, p = 0.033, all-cause mortality 7.0% vs. 1.0%, p < 0.001, and UC-related mortality 1.9% vs. 0.2%, p = 0.017 than non-elderly-onset patients.
Conclusions:
Elderly-onset UC patients are increasing in number. These patients have higher risk of opportunistic infections, hospitalisation, colorectal cancer, and mortality than non-elderly-onset patients. Management and therapeutic strategies in this special group need careful attention.
Cancer stem-like cells, possessing “stemness” properties, play crucial roles in progression, metastasis, and drug resistance in various cancers. Viral microRNAs (such as EBV-miR-BART7-3p), as ...exogenous regulators, have been discovered to regulate malignant progression of nasopharyngeal carcinoma (NPC), suggesting a possible role of viral microRNAs in imposing stemness. In this study, we found that EBV-miR-BART7-3p induce stemness of NPC cells. We firstly reported that EBV-miR-BART7-3p increased the percentage of side population cells, the development of tumor spheres, and the expression level of stemness markers
in vitro
. This viral microRNA also enhanced stem-like or cancer-initiating properties of NPC cells
in vivo
. Besides, we identified SMAD7 as a novel target gene of EBV-miR-BART7-3p in addition to PTEN gene we previously reported; this viral microRNA suppressed SMAD7, led to activation of TGF-β signaling, and eventually enhanced the stemness of NPC cells. Silencing of SMAD7 resembled the effects generated by EBV-miR-BART7-3p in NPC cells. After reconstitution of SMAD7, EBV-miR-BART7-3p-expressing cells underwent a phenotypic reversion. EBV-positive NPC cells were used to enable experimental validation. Finally, we further discovered that EBV-miR-BART7-3p increased chemo-resistance of NPC
in vitro
and
in vivo
, supporting that EBV-miR-BART7-3 resulted in increased stemness of NPC cells and lead to drug resistance and cancer recurrence. Overall, this study uncovered a novel mechanism underlying viral microRNA-associated stemness of NPC cells. This viral microRNA and its associated cellular genes may be potential therapeutic targets for restraining chemo-resistance and recurrence of NPC.
Abstract
Background
Elderly-onset inflammatory bowel disease IBD, defined as age ≥60 at diagnosis, is increasing worldwide. We aimed to compare clinical characteristics and natural history of ...elderly-onset IBD patients with those of adult-onset IBD patients.
Methods
Patients with a confirmed diagnosis of IBD from 1981 to 2016 were identified from a territory-wide Hong Kong IBD registry involving 13 hospitals. Demographics, comorbidities, clinical features, and outcomes of elderly-onset IBD patients were compared with those of adult-onset IBD patients.
Results
A total of 2413 patients were identified, of whom 270 11.2% had elderly-onset IBD. Median follow-up duration was 111 months (interquartile range IQR: 68–165 months). Ratio of ulcerative colitis UC: Crohn’s disease CD was higher in elderly-onset IBD than in adult-onset IBD patients 3.82:1 vs 1.39:1; p <0.001. Elderly-onset CD had less perianal involvement 5.4% vs 25.4%; p <0.001 than adult-onset CD. Elderly-onset IBD patients had significantly lower cumulative use of immunomodulators p = 0.001 and biologics p = 0.04. Elderly-onset IBD was associated with higher risks of: cytomegalovirus colitis (odds ratio OR: 3.07; 95% confidence interval CI 1.92–4.89; p <0.001); herpes zoster OR: 2.42; 95% CI 1.22–4.80; p = 0.12; and all cancer development hazard ratio: 2.97; 95% CI 1.84–4.79; p <0.001. They also had increased number of overall hospitalisations OR: 1.14; 95% CI 1.09–1.20; p <0.001, infections-related hospitalisation OR: 1.87; 95% CI 1.47–2.38; p <0.001, and IBD-related hospitalisation OR: 1.09; 95% CI 1.04- 1.15; p = 0.001 compared with adult-onset IBD patients.
Conclusions
Elderly-onset IBD was associated with increased risk of infections and cancer development, and increased infection- and IBD-related hospitalisations. Specific therapeutic strategies to target this special population are needed.
Nasopharyngeal carcinoma (NPC) is common among southern Chinese including the ethnic Cantonese population living in Hong Kong. Epstein-Barr virus (EBV) infection is detected in all undifferentiated ...type of NPC in this endemic region. Establishment of stable and latent EBV infection in premalignant nasopharyngeal epithelial cells is an early event in NPC development and may contribute to its pathogenesis. Immortalized primary nasopharyngeal epithelial cells represent an important tool for investigation of EBV infection and its tumorigenic potential in this special type of epithelial cells. However, the limited availability and small sizes of nasopharyngeal biopsies have seriously restricted the establishment of primary nasopharyngeal epithelial cells for immortalization. A reliable and effective method to immortalize primary nasopharyngeal epithelial cells will provide unrestricted materials for EBV infection studies. An earlier study has reported that Bmi-1 expression could immortalize primary nasopharyngeal epithelial cells. However, its efficiency and actions in immortalization have not been fully characterized. Our studies showed that Bmi-1 expression alone has limited ability to immortalize primary nasopharyngeal epithelial cells and additional events are often required for its immortalization action. We have identified some of the key events associated with the immortalization of primary nasopharyngeal epithelial cells. Efficient immortalization of nasopharyngeal epithelial cells could be reproducibly and efficiently achieved by the combined actions of Bmi-1 expression, activation of telomerase and silencing of p16 gene. Activation of MAPK signaling and gene expression downstream of Bmi-1 were detected in the immortalized nasopharyngeal epithelial cells and may play a role in immortalization. Furthermore, these newly immortalized nasopharyngeal epithelial cells are susceptible to EBV infection and supported a type II latent EBV infection program characteristic of EBV-infected nasopharyngeal carcinoma. The establishment of an efficient method to immortalize primary nasopharyngeal epithelial cells will facilitate the investigation into the role of EBV infection in pathogenesis of nasopharyngeal carcinoma.
Epstein‐Barr virus (EBV) infection has been postulated to be an early event involved in the pathogenesis of nasopharyngeal carcinomas (NPC). The lack of representative premalignant nasopharyngeal ...epithelial cell system for EBV infection has hampered research investigation into the regulation and involvement of EBV infection in NPC pathogenesis. We have compared the efficiency of EBV infection in nasopharyngeal epithelial cells with different biological properties including immortalized, primary and cancerous nasopharyngeal epithelial cells. EBV infection could be achieved in all the nasopharyngeal epithelial cells examined with variable infection rate. TGF‐β effectively enhanced EBV infection into nasopharyngeal epithelial cells both in the immortalized and primary nasopharyngeal epithelial cells. Stable infection of EBV was achieved in a telomerase‐immortalized nasopharyngeal epithelial cell line, NP460hTert. The expression pattern of EBV‐encoded genes and biological properties of this EBV infected cell line on long‐term propagation were monitored. The EBV‐infected nasopharyngeal epithelial cells acquired anchorage‐independent growth and exhibited invasive growth properties on prolonged propagation. A distinguished feature of this EBV‐infected nasopharyngeal epithelial cell model was its enhanced ability to survive under growth factor and nutrient starvation. This was evidenced by the suppressed activation of apoptotic markers and sustained activation of pAkt of EBV‐infected cells compared to control cells under nutrient starvation. Examination of cytokine profiles of EBV‐infected NP460hTert cells to nutrient and growth factor deprivation revealed upregulation of expression of MCP‐1 and GRO‐α. The establishment of a stable EBV infection model of premalignant nasopharyngeal epithelial cells will facilitate research investigation into the pathogenic role of EBV in NPC development.
Hepatocellular carcinoma (HCC) is an invasive cancer with a high rate of recurrence and metastasis. Agents with anti-proliferative as well as anti-metastatic activity will be ideal for effective ...treatment. Here, we demonstrated that berberine, an isoquinoline alkaloid, harbored potent anti-metastatic and anti-proliferative activities in vivo. Using an orthotopic model of HCC (MHCC-97L), which spontaneously develops lung metastases (one of the most common sites of HCC metastasis), we found that berberine treatment (10mg/kg/2days) significantly reduced lung metastasis from the liver tumors by ~85% (quantitated by bioluminescence emitted from lung metastases). Histological examination also confirmed the reduced incidence and number of lung metastases in berberine-treated mice. Furthermore, berberine effectively suppressed extra-tumor invasion of the primary HCC implant into the surrounding normal liver tissue, illustrating its potent anti-metastatic action in vivo. Consistent with previous reports in other cancer, berberine's anti-tumor activity was accompanied by suppression of cellular proliferation, invasiveness and HIF-1α/VEGF signaling. Strikingly, further mechanistic investigation revealed that berberine exerted profound inhibitory effect on the expression of Id-1, which is a key regulator for HCC development and metastasis. Berberine could suppress the transcription level of Id-1 through inhibiting its promotor activity. Specific downregulation of Id-1 by knocking down its RNA transcripts in HCC cells inhibited cellular growth, invasion and VEGF secretion, demonstrating the functional relevance of Id-1 downregulation induced by berberine. Lastly, berberine's anti-proliferative and anti-invasive activities could be partially rescued by Id-1 overexpression in HCC models, revealing a novel anti-cancer/anti-invasive mechanism of berberine via Id-1 suppression.
•HCC growth and metastasis are suppressed by berberine in orthotopic mice model.•Berberine reduces local invasion and lung metastasis from the liver tumors.•Berberine downregulates Id-1 expression by inhibiting the Id-1 promoter.•Id-1 knockdown inhibits cellular proliferation and invasion of HCC cells.•The anti-growth and anti-metastatic effects of berberine are mediated by Id-1.
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