Design of wormlike automated robotic endoscope: dynamic interaction between endoscopic balloon and surrounding tissues.

BACKGROUND: The current design of capsule endoscope is limited by the inability to control the motion within gastrointestinal tract. The rising incidence of gastrointestinal cancers urged improvement in the method of screening endoscopy. OBJECTIVES: This preclinical study aimed to design and develop a novel locomotive module for capsule endoscope. We investigated the feasibility and physical properties of this newly designed caterpillar-like capsule endoscope with a view to enhancing screening endoscopy. DESIGN: This study consisted of preclinical design and experimental testing on the feasibility of automated locomotion for a prototype caterpillar endoscope. The movement was examined first in the PVC tube and then in porcine intestine. The image captured was transmitted to handheld device to confirm the control of movement. The balloon pressure and volume as well as the contact force between the balloon and surroundings were measured when the balloon was inflated inside (1) a hard PVC tube, (2) a soft PVC tube, (3) muscular sites of porcine colons and (4) less muscular sites of porcine colons. RESULTS: The prototype caterpillar endoscope was able to move inward and backward within the PVC tubing and porcine intestine. Images were able to be captured from the capsule endoscope attached and being observed with a handheld device. Using the onset of a contact force as indication of the buildup of the gripping force between the balloon and the lumen walls, it is concluded from the results of this study that the rate of change in balloon pressure and volume is two good estimators to optimize the inflation of the balloon. CONCLUSION: The results of this study will facilitate further refinement in the design of caterpillar robotic endoscope to move inside the GI tract.

Susceptibility of Hep3B cells in different phases of cell cycle to tBid.

tBid is a pro-apoptotic molecule. Apoptosis inducers usually act in a cell cycle-specific fashion. The aim of this study was to elucidate whether effect of tBid on hepatocellular carcinoma (HCC) Hep3B cells was cell cycle phase specific. We synchronized Hep3B cells at G0/G1, S or G2/M phases by chemicals or flow sorting and tested the susceptibility of the cells to recombinant tBid. Cell viability was measured by MTT assay and apoptosis by TUNEL. The results revealed that tBid primarily targeted the cells at G0/G1 phase of cell cycle, and it also increased the cells at the G2/M phase. 5-Fluorouracil (5-FU), on the other hand, arrested Hep3B cells at the G0/G1 phase, but significantly reduced cells at G2/M phase. The levels of cell cycle-related proteins and caspases were altered in line with the change in the cell cycle. The combination of tBid with 5-FU caused more cells to be apoptotic than either agent alone. Therefore, the complementary effect of tBid and 5-FU on different phases of the cell cycle may explain their synergistric effect on Hep3B cells. The elucidation of the phase-specific effect of tBid points to a possible therapeutic option that combines different phase specific agents to overcome resistance of HCC.

Induction of autophagy in hepatocellular carcinoma cells by SB203580 requires activation of AMPK and DAPK but not p38 MAPK.

SB203580 is a well-known inhibitor of p38 mitogen-activated protein kinase (MAPK). However, it can suppress cell proliferation in a p38 MAPK independent manner. The inhibitory mechanism remains unknown. Here, we showed that SB203580 induced autophagy in human hepatocellular carcinoma (HCC) cells. SB203580 increased GFP-LC3-positive cells with GFP-LC3 dots, induced accumulation of autophagosomes, and elevated the levels of microtubule-associated protein light chain 3 and Beclin 1. It stimulated the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) and p53, but inhibited the phosphorylation of death-associated protein kinase (DAPK). Inhibition of AMPK, p53, or DAPK attenuated SB203580-induced autophagy. AMPK activation appeared to predate the DAPK signal. The activation of both AMPK and DAPK prompted the phosphorylation of p53 and enhanced Beclin 1 expression. Neither the downregulation of p38 MAPK by its siRNA or chemical inhibitor nor the upregulation of p38 MAPK by p38 MAPK DNA transfection affected B203580-induced autophagy. Collectively, the findings demonstrate a novel function of SB203580 to induce autophagy via activating AMPK and DAPK but independent of p38 MAPK. The induction of autophagy can thus account for the antiproliferative effect of SB203580 in HCC cells.

Ent-11α-hydroxy-15-oxo-kaur-16-en-19-oic-acid inhibits hepatocellular carcinoma in vitro and in vivo via stabilizing IkBα.

Ent-11-hydroxy-15-oxo-kaur-16-en-19-oic-acid (5F) isolated from Pteris Semipinnata L is known to inhibit certain tumor cells in vitro. The information on the in vivo effect of 5F is limited and its effect on hepatocellular carcinoma (HCC) is unknown. In this study, the anti-tumor effect of 5F was investigated in a diethylnitrosamine (DEN)-induced mouse HCC model. In addition to therapeutic effect, the potential side effect was monitored. A panel of cultured HCC cells was used to confirm the in vivo data and explore the responsible molecular pathway. The result showed that 5F significantly inhibited the DEN-induced HCC tumors by reducing the number of tumor foci and the volume of tumors. Furthermore, 5F induced the death of cultured HCC cells in dose- and time-dependent manners. The cell death was confirmed to be apoptotic by in vivo and in vitro TUNEL assays. 5F inhibited NF-kB by stabilizing its inhibitor IkBα, reducing the nuclear p65 and inhibiting NF-kB activity. Subsequently it affected the NF-kB downstream molecules with a decrease in anti-apoptotic Bcl-2 and increase in pro-apoptotic Bax and Bak. During the whole period of the experiment, mice receiving 5F appeared to be healthy, though they suffered from a mild degree of hair loss. 5F did not damage liver and renal functions. In conclusion, 5F is effective against HCC with minimal side effects. It induces apoptosis in HCC cells via inhibiting NF-kB, leading to the decrease of Bcl-2 but the increase of Bax and Bak.

Thromboxane synthase suppression induces lung cancer cell apoptosis via inhibiting NF-κB.

Accumulating evidence shows that the inhibition of thromboxane synthase (TXS) induced apoptosis in cancer cells. TXS inhibitor 1-Benzylimidzole (1-BI) can trigger apoptosis in lung cancer cells but the mechanism is not fully defined. In this study, lung cancer cells were treated with 1-BI. In this study, the level of reactive oxygen species (ROS) was measured and NF-κB activity was determined in human lung cancer cells. The roles of ROS and NF-κB in 1-BI-mediated cell death were analyzed. The results showed that 1-BI induced ROS generation but decreased the activity of NF-κB by reducing phosphorylated IκBα (p-IκBα) and inhibiting the translocation of p65 into the nucleus. In contrast to 1-BI, antioxidant N-acetyl cysteine (NAC) stimulated cell proliferation and significantly protected the cells from 1-BI-mediated cell death by neutralizing ROS. Collectively, apoptosis induced by 1-BI is associated with the over-production of ROS and the reduction of NF-κB. Antioxidants can significantly block the inhibitory effect of 1-BI.

Inhibition of thromboxane synthase induces lung cancer cell death via increasing the nuclear p27.

The role of thromboxane in lung carcinogenesis is not clearly known, though thromboxane B2 (TXB(2)) level is increased and antagonists of thromboxane receptors or TXA2 can induce apoptosis of lung cancer cells. p27, an atypical tumor suppressor, is normally sequestered in the nucleus. The increased nuclear p27 may result in apoptosis of tumor cells. We hypothesize that the inhibition of thromboxane synthase (TXS) induces the death of lung cancer cells and that such inhibition is associated with the nuclear p27 level. Our experiment showed that the inhibition of TXS significantly induced the death or apoptosis in lung cancer cells. The activity of TXS was increased in lung cancer. The nuclear p27 was remarkably reduced in lung cancer tissues. The inhibition of TXS caused the cell death and apoptosis of lung cancer cells, likely via the elevation of the nuclear p27 since the TXS inhibition promoted the nuclear p27 level and the inhibition of p27 by its siRNA recovered the cell death induced by TXS inhibition. Collectively, lung cancer cells produce high levels of TXB(2) but their nuclear p27 is markedly reduced. The inhibition of TXS results in the p27-related induction of cell death in lung cancer cells.

Keeping the Traction on in Orthopaedics.

The trauma and orthopaedic speciality continues to advance as surgery becomes more accessible and safe. However, the bygone days of treatment with traction still has its merits and should remain a part of practitioner's repertoire. This will allow the practitioners to be resourceful in times of unexpected scenarios. We aim to write this article to describe indications, applications of various forms of traction, and their relevant complications.

Modernising Anatomy Teaching: Which Resources Do Students Rely On?

The way in which we learn anatomy has changed exponentially over the decades and students now have access to lecture notes, textbooks, computer-assisted programmes, and a wide variety of internet based information. This study explored which resources were the most (and least) useful for a group of first year, undergraduate, medical students, with minimal prior content exposure (aged 18 and 19 years old, n = 76), over an 18 month period. Anatomy websites were found to be the most useful (30%), followed by tutorials (20%) and lectures (19%). A total of 13% found the university computer-assisted learning (CAL) platform least useful. We subsequently enhanced our 'urogenital' CAL anatomy module, with inclusion of new and updated images, videos and tutorials, as well as, digital and printed 3D-models. A post-intervention survey (n = 81) showed an increase from 12% to 27% for CAL as being most useful, and a decrease from 13% to 3% as being least useful. Our results provided a snapshot of students' preferences in studying anatomy, and highlighted the importance of digital platforms and the need for evaluating our own learning resources. We must be mindful that there is an increasing tendency for students to rely on the Internet for information, which may expose them to unfiltered and unreliable content. We conclude that educators must be aware of the spectrum of learning resources used by students, to ensure that our own Institutional eLearning platforms are optimised to meet the diverse needs of learners.

Conservative approach to the acute management of a large mesenteric cyst.

Mesenteric cysts are rare, benign gastrointestinal cystic lesions, which are often non-troublesome and present as an incidental radiological finding. However, surgery is often performed in the acute setting to remove lesions that are symptomatic. This report highlights the case of a large, symptomatic mesenteric cyst managed successfully with initial conservative measures followed by planned elective surgery. A 44-year-old female presented with a four-day history of generalised abdominal pain associated with distension, fever, diarrhoea and vomiting. Computer tomography revealed a large (21.7 cm × 11.8 cm × 14 cm) mesenteric cyst within the left abdomen cavity. She was admitted and treated conservatively with intravenous fluids and antibiotics for four days, which lead to complete symptom resolution. Follow-up at intervals of one and three months revealed no return of symptoms. An elective laparotomy and excision of the mesenteric cyst was then scheduled and performed safely at nine months after the initial presentation. Compared to acute surgery, acute conservative management followed by planned elective resection of a symptomatic mesenteric cyst may prove safer. The withholding of an immediate operation may potentially avoid unnecessary operative risk and should be considered in patients without obstructive and peritonitic symptoms. Our case demonstrated the safe use of initial conservative management followed by planned elective surgery of a mesenteric cyst found in the acute setting, which was symptomatic but was not obstructive or causing peritonitic symptoms.

A Therapeutic Wireless Capsule for Treatment of Gastrointestinal Haemorrhage by Balloon Tamponade Effect.

OBJECTIVE: Wireless capsule endoscope (WCE) is a revolutionary approach to diagnose small bowel pathologies. Currently available WCEs are mostly passive devices with image capturing function only, while on-going efforts have been placed on robotizing WCEs or to enhance them with therapeutic functions. In this paper, the authors present a novel inflatable WCE for haemostasis in the gastrointestinal (GI) tracts by balloon tamponade effect. METHODS: The proposed wireless capsule consists of a balloon that can be inflated using the endothermic reaction of acid and base. When the balloon reached a precalculated pressure level, it is able to stop at a bleeding site in the bowel, and achieve haemostasis by tamponade effect. The prototype is 14 mm in diameter, with three sections of 13, 35, and 12 mm in length, respectively. The three sections are linked together with flexible joints and enclosed in a silicone balloon. The prototypes were tested in ex vivo porcine intestine models. RESULTS: In the ten ex vivo trials conducted, the inflatable wireless capsule achieved average balloon pressure of 46.0 mmHg and withstood average maximum longitudinal pulling force at 1.46 N. An in vivo study was carried out as a proof-of-concept for treating bleeding in a porcine model. The proposed inflatable WCE succeeded in the animal test by controlling haemostasis within 5 min. No rebleeding was observed in the next 20 min. CONCLUSION: The results suggested that the inflatable capsule with a real-time bleeding detection algorithm can be implemented. Moreover, the proposed inflatable WCE prototype can achieve haemorrhage control in the lower GI. SIGNIFICANCE: To our best knowledge, this is the first study that demonstrated the potential to treat GI haemorrhage by an inflatable WCE. The proposed capsule enables the development of a closed-loop system based on a body sensor network to provide early treatment of GI bleeding for p-medicine.

Cytochrome P450 1A2 Metabolizes 17ß-Estradiol to Suppress Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) occurs more frequently in men than in women. It is commonly agreed that estrogen plays important roles in suppressing HCC development, however, the underlying mechanism remains largely unknown. Since estrogen is mainly metabolized in liver and its metabolites affect cell proliferation, we sought to investigate if the liver-specific cytochrome P450 1A2 (CYP1A2) mediated the inhibitory effect of estrogen on HCC. In this study, the expression of estrogen-metabolizing enzyme CYP1A2 was determined in HCC tissues and cell lines. Cell proliferation and apoptosis were assessed in cells with or without CYP1A2 overexpression. The levels of 17ß-estradiol (E2) and its metabolite 2-methoxyestradiol (2-ME) were determined. A xenograft tumor model in mice was established to confirm the findings. It was found that CYP1A2 expression was greatly repressed in HCC. E2 suppressed HCC cell proliferation and xenograft tumor development by inducing apoptosis. The inhibitory effect was significantly enhanced in cells with CYP1A2 overexpression, which effectively conversed E2 to the cytotoxic 2-ME. E2 in combination with sorafenib showed an additive effect on HCC. The anti-HCC effect of E2 was not associated with estrogen receptors ERα and ERß as well as tumor suppressor P53 but enhanced by the approved anti-HCC drug sorafenib. In addition, HDAC inhibitors greatly induced CYP1A2 promoter activities in cancer cells, especially liver cancer cells, but not in non-tumorigenic cells. Collectively, CYP1A2 metabolizes E2 to generate the potent anti-tumor agent 2-ME in HCC. The reduction of CYP1A2 significantly disrupts this metabolic pathway, contributing the progression and growth of HCC and the gender disparity of this malignancy.

Cell death induced by Pteris semipinnata L. is associated with p53 and oxidant stress in gastric cancer cells.

In this study, we demonstrated that Ent-11alpha-hydroxy-15-oxo-kaur-16-en-19-oic-acid (5F) had stronger cytotoxicity against MKN-45, a gastric cancer cell line bearing wild-type p53 than MKN-28, another gastric cancer cell line containing missense mutation in p53. The rapid increase of ROS level was involved in the mechanism of cytotoxicity. Classical features of apoptosis induced by 5F were observed in MKN-45 cells only or more significant in MKN-45 cells than MKN-28 cells. Translocation of Bax from cytosol to mitochondria, reduction of delta psi m and DNA fragmentation were induced by 5F in the p53-dependent manner. We conclude that the expression of Bax and its downstream molecules requires the presentation of a wild-type p53 in the cells treated by 5F.

A mucoadhesive endoluminal wearable sensory system.

Bio- or muco-adhesive anchoring is a challenge for the development of advanced gastrointestinal (GI) surgical instruments, endoluminal monitoring devices and drug delivery systems. In this paper, we present a polymeric bio-adhesive film embedded with an optical sensor that can potentially be used to detect gastrointestinal bleeding. Four different formulas of mucoadhesive polymers were synthesized based on various chemical components and concentration combinations, and they were further layered with miniature photoplethymographic (PPG) sensors. The adhesive ability of the proposed mucoadhesive-sensor module was tested by attaching it to the lumen of a porcine stomach and compared amongst the four formulas. pH testing was also implemented to simulate the performance of the film in gastric cavity. To demonstrate the signal quality of this module, we also tested on the skin of five healthy subjects for hours. The observed shear detachment force between mucoadhesive film and porcine stomach tissue of all four formulations ranged from 0.09 to 1.38 N, and the performance of mucoadhesive film in pH 7 and pH 2 were similar. The module can attach firmly onto the skin for 3-10 hours with comparable PPG signal quality to traditional clip-based setup. With the advent of mucosal tissue anchoring by means of bioadhensive film, a wider extent of endoluminal procedures may become feasible. This emerging technology can also help shape the future of in-body wearable devices in the GI tract or other endoluminal cavities.

A randomized controlled trial of nasolaryngoscopy training techniques.

OBJECTIVES/HYPOTHESIS: Flexible nasolaryngoscopy is an essential skill for otolaryngology trainees to develop, but there is a lack of standardized training for this procedure. The aim of this study was to assess whether using training on a realistic human mannequin together with structured video feedback improved trainees' performance at flexible nasolaryngoscopy. STUDY DESIGN: Three-armed, single-blinded, randomized controlled study. METHODS: Thirty-six junior doctors and final-year medical students were randomly allocated to one of three groups. All received a lecture and video presentation on flexible nasolaryngoscopy. One group received additional tuition using a training mannequin. The last group received mannequin training and feedback on their performance using a video recording. The trainees then undertook flexible nasolaryngoscopy on volunteers with these endoscopies recorded. Blinded observers scored the trainees on a range of objective and subjective measures. The volunteers who were also blinded to the candidates' training scored the comfort of the procedure. RESULTS: Adding mannequin training showed a trend toward improvement of performance but did not reach statistical significance. Mannequin training together with video feedback produced significant performance improvement in patient comfort (P = .0065), time to reach the vocal folds (P = .017), and global ability (P = .0006). Inter-rater reliability was excellent with P < .01 in all assessments. CONCLUSIONS: Simulation-based training using an anatomically correct model of the upper airway together with formalized video-assisted feedback on that training is a simple and effective way to improve endoscopy skills prior to starting flexible nasolaryngoscopy on patients.

A study of the combination of triamcinolone and 5-fluorouracil in modulating keloid fibroblasts in vitro.

BACKGROUND: Preliminary clinical trials suggest a superior effect when steroids and 5-fluorouracil are injected together for the intralesional therapy of keloids. In addition, it has been proposed that low-dose 5-fluorouracil may have advantages over conventional high dosages. We explored the molecular basis for the potential synergy involved in the combined treatment with triamcinolone and 5-fluorouracil. METHODS: The effects of triamcinolone alone or in combination with low-dose 5-fluorouracil on cell proliferation, cell-cycle progression, apoptosis and regulation of p53, p21, type I collagen (Col-1), vascular endothelial growth factor (VEGF), transforming growth factor-beta1 (TGF-beta1) and matrix metalloproteinase-2 (MMP-2) production in primary cultured keloid fibroblasts were examined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), flow cytometric and Western blotting assays. RESULTS: Triamcinolone suppressed cell proliferation and induced G1 cell-cycle arrest but not apoptosis. By contrast, 5-fluorouracil induced G2 cell-cycle arrest and apoptosis that may be associated with p53 activation and p21 up-regulation. The modulation of Col-1, VEGF, TGF-beta1 and MMP-2 expression by triamcinolone and 5-fluorouracil alone or their combination varied between individual cell lines; the trend is to promote a reduction in Col-1 and TGF-beta1 but up-regulation of MMP-2 expression. 5-Fluorouracil played a predominant role in the combined treatment leading to more significant cell proliferation inhibition, apoptosis, Col-1 suppression and MMP-2 induction (p < 0.05). CONCLUSIONS: Our data provide the molecular-based evidence for the observed clinical benefits of adding 5-fluorouracil to a steroid injection for improved scar regression and reduced recurrence of keloids. We expect fewer undesirable side effects in the combined treatment when the lower therapeutic dose of the individual drugs is to be used.

A high-sensitivity near-infrared phototransistor based on an organic bulk heterojunction.

High-gain photodetectors with near-infrared (NIR) sensitivity are critical for biomedical applications such as photoplethysmography and optical coherence tomography where detected optical signals are relatively weak. Current photodetection technologies rely on avalanche photodiodes and photomultipliers to achieve high sensitivity. These devices, however, require a high operation voltage and are not compatible with CMOS based read-out circuits (ROCs). In this work we demonstrate a solution-proceeded NIR phototransistor structure based on a bulk heterojunction (BHJ) of a narrow bandgap polymer, poly(N-alkyl diketopyrrolo-pyrrole dithienylthieno[3,2-b]thiophene) (DPP-DTT), and [6,6]-phenyl-C61-butyric acid methylester (PCBM). The device exhibits ultrahigh responsivity (∼5 × 10(5) A W(-1)) as well as wide tunability (>1 × 10(4)) of photoconductive gain. Using the current-voltage and transient photocurrent measurements we show that the high responsivity is due to the combined effects of fast transport of holes in the polymer matrix and slow detrapping of electrons from the isolated PCBM domains. The wide gain tunability and the efficient suppression of noise current are achieved through the use of the optically tunable gate terminal. We demonstrate that our phototransistor can be used as the detection unit in a photoplethysmography sensor for non-invasive, continuous finger pulse wave monitoring. The high-sensitivity of the phototransistor allows the use of a low-power light source, thus reducing the overall power consumption of the sensor. This, together with the solution processibility and the simple device configuration (which is compatible with conventional ROCs), make the phototransistor a very promising component for the next generation low-cost, mobile biomedical devices for health monitoring and remote diagnostics.

A clip-free eyeglasses-based wearable monitoring device for measuring photoplethysmograhic signals.

An eyeglasses-based device has been developed in this work to acquire photoplethysmogram (PPG) from the nose bridge. This device is aimed to provide wearable physiological monitoring without uncomfortable clips frequently used in PPG measurement from finger and ear. Switching control is applied on the LED and photo detector for power saving. An experiment involving postural change and treadmill jogging among 10 healthy young subjects was carried out to evaluate the performance of the device. Electrocardiogram (ECG) and PPG from finger, ear and nose were simultaneously recorded, from which heart rate (HR) and pulse transit time (PTT) were calculated. The results show that PPG measured from nose and ear are more resistant to motion than signal from finger during exercise. In addition, the difference between PTT measured from ear and nose indicates that local vasomotor activities may exist on ear and/or nose channel, and suggests that PPG from different sites should be used for cuff-less PTT-based BP estimation. We conclude that this wearable device has great potential to be used in the healthcare management in the future.

Differential functions of C- and N-terminal hepatitis B x protein in liver cells treated with doxorubicin in normoxic or hypoxic condition.

Hepatitis viral B x protein (HBx), a hepatocarcinogen, is frequently mutated. Hypoxia influences the growth of HCC and also the sensitivity of tumor cells to treatments. We aimed to test the role of HBx and acute hypoxia in the efficacy of chemotherapy. In this study, we established 4 Chang liver cell lines with the full-length HBx (HBx), the first 50 amino acids of N-terminal HBx (HBx/50), the last 104 amino acids of C-terminal HBx (HBx/51) and empty vector (CL), respectively. MTT and TNUEL assays were used to assess cell viability and apoptosis respectively. Western blot was used to determine the expression of relevant proteins. Results showed that among 4 cell lines, doxorubicin was most effective in decreasing the viability and enhancing apoptosis in HBx/51 cells, while HBx/50 cells were most resistant to the treatment. Cells in hypoxia were more susceptible to doxorubicin than cells in normoxia. Hypoxia facilitated the Bid cleavage especially in HBx/51 cells via phosphorylating p38 MAPK. p38 MAPK inhibitor significantly reduced the tBid level and increased cell viability. In conclusion, N-terminal HBx and C-terminal HBx function differentially in their ability to regulate cell growth, with the former being promotive but the latter being inhibitory. The acute hypoxia may overcome the HBx-induced resistance and facilitate the chemotherapy.

5-fluorouracil mediates apoptosis and G1/S arrest in laryngeal squamous cell carcinoma via a p53-independent pathway.

PURPOSE: 5-Fluorouracil (5-FU) is a commonly used chemotherapeutic agent in the treatment of laryngeal squamous cell carcinoma. 5-FU can induce cell cycle arrest or apoptosis in various cancers via either a p53-dependent or a p53-independent pathway; however, its pathway of action in laryngeal carcinoma is unknown. In this study, we aim to investigate the role that p53 plays in the cytotoxic effect of 5-FU on laryngeal squamous carcinoma cells. MATERIALS AND METHODS: We employed two human laryngeal squamous carcinoma cell lines with different p53 statuses-one (UMSCC12) had truncated non-functional p53 and the other (UMSCC11A) had mutant but functional p53. Cell death was detected using cytotoxicity assay and Annexin V staining. Cell cycles were analyzed by flow cytometry. Western blot was used to analyze the protein expression. RESULTS: 5-FU induces apoptosis in both UMSCC12 and UMSCC11A cells in a dose- and time-dependent manner, suggesting that the pathway was p53-independent. 5-FU induced the accumulation of retinoblastoma protein and a cyclin dependent kinase inhibitor, p21WAF1/CIP1, in both UMSCC12 and UMSCC11A cells. However, 5-FU did not induce p53 expression in either UMSCC12 or UMSCC11A cells. In addition, G1/S cell cycle phase arrest was associated with antiproliferative activity of 5-FU in both cell lines. In order to gain an insight into the role p53 plays in response to 5-FU treatment in laryngeal carcinoma, we further transfected either a wildtype p53 plasmid or an empty pcDNA3.1 vector into UMSCC12 cells. We found that 5-FU increased pRb and p21WAF1/CIP1 expression in both p53-transfected and vector-transfected cells without the significant accumulation of p53. DISCUSSION: Our results suggest that 5-FU mediates apoptosis and G1/S cell cycle phase arrest in laryngeal carcinoma via a p53-independent but p21WAF1/CIP1-dependent or p21WAF1/CIP1-Rb-dependent pathway. While p53 does not seem to be involved in 5-FU induced apoptosis and cell cycle arrest in laryngeal carcinoma, further studies are needed to examine the roles of retinoblastoma protein and p21WAF1/CIP1 in laryngeal carcinoma receiving chemotherapy.

Glioblastoma cells deficient in DNA-dependent protein kinase are resistant to cell death.

DNA-dependent protein kinase (DNA-PK), a nuclear serine/threonine kinase, is responsible for the DNA double-strand break repair. Cells lacking or with dysfunctional DNA-PK are often associated with mis-repair, chromosome aberrations, and complex exchanges, all of which are known to contribute to the development of human cancers including glioblastoma. Two human glioblastoma cell lines were used in the experiment, M059J cells lacking the catalytic subunit of DNA-PK, and their isogenic but DNA-PK proficient counterpart, M059K. We found that M059K cells were much more sensitive to staurosporine (STS) treatment than M059J cells, as demonstrated by MTT assay, TUNEL detection, and annexin-V and propidium iodide (PI) staining. A possible mechanism responsible for the different sensitivity in these two cell lines was explored by the examination of Bcl-2, Bax, Bak, and Fas. The cell death stimulus increased anti-apoptotic Bcl-2 and decreased pro-apoptotic Bcl-2 members (Bak and Bax) and Fas in glioblastoma cells deficient in DNA-PK. Activation of DNA-PK is known to promote cell death of human tumor cells via modulation of p53, which can down-regulate the anti-apoptotic Bcl-2 member proteins, induce pro-apoptotic Bcl-2 family members and promote a Bax-Bak interaction. Our experiment also demonstrated that the mode of glioblastoma cell death induced by STS consisted of both apoptosis and necrosis and the percentage of cell death in both modes was similar in glioblastoma cell lines either lacking DNA-PK or containing intact DNA-PK. Taken together, our findings suggest that DNA-PK has a positive role in the regulation of apoptosis in human glioblastomas. The aberrant expression of Bcl-2 family members and Fas was, at least in part, responsible for decreased sensitivity of DNA-PK deficient glioblastoma cells to cell death stimuli.