Identification of measurable nontransient responses to low-dose radiation in human primary cell cultures remains a problem. To this end, circulating endothelial colony-forming (progenitor) cells (ECFCs) were examined as an experimental model. ECFCs were isolated from three cord blood donors. Cells were positive for endothelial cell markers and remained highly proliferative after long-term cryopreservation. A single dose of X-ray radiation (0.06–0.38 Gy) inhibited ECFC culture growth. This effect was evident at 48 hours and persisted up to 72 hr postirradiation. Such protracted cytostatic response of ECFCs to low-dose radiation suggests that ECFC primary cultures can be used to study low-dose radiation effects.
Cancer biomarkers; Cancer prevention; Carcinogenesis
A 27-year-old man with progressive bilateral visual decline was diagnosed to have Bietti's crystalline dystrophy (BCD). Fluorescein angiography revealed bilateral petaloid type late hyperfluorescence implicating concurrent cystoid macular edema (CME). Optical coherence tomography exhibited cystoid foveal lacunas OU. During the follow-up of six years, intraretinal crystals reduced in amount but CME persisted angiographically and tomographically. CME is among the rare macular features of BCD including subfoveal sensorial detachment, subretinal neovascular membrane, and macular hole.
Patient: Female, 16
Final Diagnosis: Malrotation and cholecystoduodenal fistula
Symptoms: Abdominal pain • anorexia • fever • nausea • vomiting
Clinical Procedure: —
Specialty: Gastroenterology and Hepatology
Cholecystoduodenal fistula (CDF) is the most common cholecystenteric fistula. It is a late complication of gallbladder disease with calculus and is mainly encountered in the elderly and females.
We report the case of a teenage patient with cholecystoduodenal fistula and malrotation. Direct plain abdominal x-ray demonstrated air in the biliary system. Computed tomography revealed CDF-associated with an anomaly of intestinal malrotation. She had gallstones (with a few stones in the gallbladder) and cholecystitis. CDF is caused by malrotation, and cholecystitis has not been reported before. In this regard our patient is the first and youngest reported case.
We suggest that CDF is probably a consequence of malrotation. The patient’s clinical features and operative management are presented and discussed with current literature.
cholecystoduodenal fistula; cholecystitis; malrotation
Small ubiquitin-like modifier (SUMO1, 2, 3) is a group of proteins that conjugate to lysine residues of target proteins thereby modifying their activity, stability, and subcellular localization. A large number of SUMO target proteins are transcription factors and other nuclear proteins involved in gene expression. Furthermore, SUMO conjugation plays key roles in genome stability, quality control of newly synthesized proteins, proteasomal degradation of proteins and DNA damage repair. Any marked increase in levels of SUMO-conjugated proteins is therefore expected to have a major impact on the fate of cells. We show here that SUMO conjugation is activated in human astrocytic brain tumors. Levels of both SUMO1- and SUMO2/3-conjugated proteins were markedly increased in tumor samples. The effect was least pronounced in low-grade astrocytoma (WHO Grade II) and most pronounced in glioblastoma multiforme (WHO Grade IV). We also found a marked rise in levels of Ubc9, the only SUMO conjugation enzyme identified so far. Blocking SUMO1-3 conjugation in glioblastoma cells by silencing their expression blocked DNA synthesis, cell growth and clonogenic survival of cells. It also resulted in DNA-PK-dependent phosphorylation of H2AX, indicative of DNA double-strand damage, and G2/M cell cycle arrest. Collectively, these findings highlight the pivotal role of SUMO conjugation in DNA damage repair processes and imply that the SUMO conjugation pathway could be a new target of therapeutic intervention aimed at increasing the sensitivity of glioblastomas to radio- and chemotherapy.
astrocytic brain tumors; DNA damage repair; DNA-PK; G2/M checkpoint; gene silencing; glioblastoma; H2AX; human; miRNA; SUMO conjugation; Ubc9
A 64-year-old male who had a macula-on superior bullous retinal detachment in OD underwent scleral buckling, 20-gauge-pars plana vitrectomy, internal drainage of subretinal fluid with perfluorocarbon fluid, 360° endolaser and perflourocarbon-fluid-air exchange surgery. Patient sat upright immediately after the surgery for the night. At the first postoperative morning although the retina was attached, there was a macular fold extending toward the temporal retinal periphery. Patient denied further surgery. During the follow-up, retinal fold gradually became less visible and it could be noticeable only by fundus autoflorescence imaging at the sixth postoperative year with a subtle epiretinal membrane formation on the optical coherence tomographic section.
We injected an intravitreal dexamethasone implant in two eyes of 2 pediatric patients with Coats’ disease in addition to other treatment modalities, such as intravitreal ranibizumab injection and indirect laser photocoagulation. In both eyes, intraocular pressure moderately rose in a temporary fashion. The dexamethasone implant seems to be a valuable addition to the armamentarium of treatment options for Coats’ disease as it necessitates fewer injections than anti-VEGF injections and thereby fewer sessions of general anesthesia in the pediatric population.
Coats’ disease; Dexamethasone implant; Laser photocoagulation; Ranibizumab
Uncontaminated urine samples are indispensable to precisely diagnose urinary tract infections in new-borns or infants. Among many clinical interventions for urine collection are described, the most common noninvasive practice is using sterile bags, associated with significant contamination of samples. In children, however, invasive methods i.e. catheterization, are generally needed for reliable urine specimens. Almost always all the inserted catheters are easily drawn back, nevertheless, might not work as expected, and lead to considerable problems that cannot be overcome. Herein, a case of a female newborn treated with a successful percutaneous suprapubic cystoscopic procedure for extracting knotted urinary catheter in the bladder is presented. The least invasive and easiest technic is suggested to be used when catheter is knotted in the bladder, as elaborately stated.
Knotted urethral catheter; urinary catheterization; percutaneous cystoscopy
In this brief report, we share our observations on a splitted Dexamethasone implant (Ozurdex) which we discovered a week after the injection. It is likely that implant splitting neither changes the efficacy of the implant nor creates a mishap for the patient.
The aim of this study was to create a diagnostic model using the artificial neural networks (ANNs) to predict malignancy in multinodular goiter patients with an indeterminate cytology.
Materials and Methods:
Out of 623 patients, 411 evaluated for multinodular goiter between July 2004 and March 2010 had a fine-needle aspiration biopsy. All patients underwent total thyroidectomy. The interpretation was consistent with an indeterminate lesion in 116 (18.6%) patients. Patient's medical records including age, sex, dominant nodule size, pre-operative serum thyroid-stimulating hormone level, thyroid hormone therapy and final pathologic diagnosis were collected retrospectively.
The mean age of the patients was 44.6 years (range, 17-78 years). About 104 (89.7%) were female and 12 (10.3%) were male patients. Final pathology revealed 24 malignant diseases (20.7%) and 92 (79.3%) benign diseases. After the completion of training, the ANN model was able to predict diagnosis of malignancy with a high degree of accuracy. The area under the curve of ANNs was 0.824.
The ANNs technique is a useful aid in diagnosing malignancy and may help reduce unnecessary thyroidectomies in multinodular goiter patients with an indeterminate cytology. Further studies are needed to construct the optimal diagnostic model and to apply it in the clinical practice.
Artificial neural network; fine-needle aspiration; ındeterminate cytology; multinodular goiter; thyroid
Sorafenib is a multi-targeted tyrosine kinase receptor inhibitor used to treat patients with advanced gastrointestinal stromal tumors (GISTs). The present study evaluated the efficacy and tolerability of sorafenib therapy for patients with GISTs. Between January 2001 and November 2012, 25 patients, from multiple centers, who had received sorafenib as the third- or fourth-line treatment for GISTs were investigated retrospectively. In total, 17 patients were male and eight were female. The median age was 54.0 years (range, 16–82 years). From the patients, 21 received imatinib for longer than six months and four received it for less than six months. The clinical benefit rate of sorafenib was 40.0%. Treatment-related adverse events were reported in 72% of patients. These adverse events were generally mild to moderate in intensity. The median progression-free survival (PFS) and overall survival (OS) times of the patients who received sorafenib were 7.2 and 15.2 months, respectively. The duration of imatinib usage was an independent prognostic factor for PFS and OS. Sorafenib is an effective treatment in patients with GISTs showing a clinical benefit rate of 40.0% and an acceptable tolerability.
sorafenib; gastrointestinal stromal tumors; efficacy
To report the efficacy of intravitreal dexamethasone implant in a patient with retinitis pigmentosa and bilateral cystoid macular edema unresponsive to topical carbonic anhydrase inhibitors.
A 36-year-old man with bilateral cystoid macular edema associated with retinitis pigmentosa that was unresponsive to topical carbonic anhydrase inhibitors underwent bilateral 0.7-mg intravitreal dexamethasone implants two weeks apart. Spectral domain optical coherence tomography revealed resolution of macular edema one week following each injection in both eyes and his visual acuity improved. However, macular edema recurred two months later in OS and three months later in OD. Second implant was considered for both eyes. No implant-related complication was experienced during the follow-up of seven months.
Inflammatory process seems to play a role in retinitis pigmentosa. Intravitreal dexamethasone implant may offer retina specialists a therapeutic option especially in cases unresponsive to other treatment regimens in eyes with retinitis pigmentosa-related macular edema.
Intravitreal dexamethasone implant; Retinitis pigmentosa; Macular edema
Use of mobile phones has rapidly risen among adolescents despite a lack of scientific certainty on their health risks. Risk perception is an important determinant of behavior, and studies on adolescents’ risk perceptions of mobile phones or base stations are very scarce. This study aims to evaluate high school students’ risk perceptions on mobile phones and base stations, their trust to authorities, their opinions regarding incivility while using mobile phones and to assess associated factors.
For this cross-sectional study, 2530 students were chosen with stratified cluster sampling among 20,493 high school students studying in Bornova district of Izmir, Turkey, among whom 2240 (88.5%) participated. Risk perceptions and opinions were questioned with a 5-point Likert scale for 24 statements grouped under four dimensions. The mean responses to the four dimensions were categorized as <3.5 (low) and ≥3.5 (high) and the determinants were analyzed with logistic regression.
Mean risk perception scores for the mobile phone, base station, trust to authority and incivility dimensions were 3.69 ± 0.89, 4.34 ± 0.78, 3.77 ± 0.93, 3.16 ± 0.93 and the prevalence of high risk perception was 65.1%, 86.7%, 66.2%, 39.7%, respectively. In the mobile phone dimension; students attending industrial technical high school had lower risk perceptions while female students, lower mothers’ education groups and students not using mobile phones (OR = 2.82, 95% CI = 1.80-4.40) had higher risk perceptions. In the base station dimension girls had higher risk perceptions (OR = 1.68, 95% CI = 1.20-2.37). Girls and students attending industrial technical high school had significantly lower risk perception however 11-12th grade group perceived the risk higher (OR = 1.45 95% CI = 1.15-1.84) in the trust to authority dimension. For the incivility dimension, female students (OR = 1.44, 95% CI = 1.14-1.82), illiterate/only literate mothers (OR = 1.79, 95% CI = 1.04-2.75) and students not using mobile phones (OR = 2.50, 95% CI = 1.62-3.87) perceived higher risk.
Understanding the effects of these determinants might aid in developing more effective educational interventions to specific subgroups on this topic. As debates on the health consequences of electromagnetic fields continue, it would be cautious to approach this issue with a preventive perspective. Efforts should be made to equalize the varying level of knowledge and to ensure that students are informed accurately.
Risk perception; Adolescent; Mobile phone; Base station; Electromagnetic field
To evaluate the effect of a single intravitreal ranibizumab injection in eyes with acute nonarteritic ischemic optic neuropathy (NAION).
Subjects and Methods:
In this retrospective clinical data analysis, 17 eyes of sixteen patients who experienced a visual loss with duration of 15 days or less comprised the study group. In addition to standard ophthalmic examination, retinal nerve fiber layer thickness (RNFLT) analysis with spectral domain OCT was also performed prior to 0.5 mg Ranibizumab injection, one week, one, three, six months and one year after the injection.
The mean time between visual loss and intravitreal injection was 7.5 days (Range, 2-15 days). Mean age of patients was 59 years (Range, 41-90 years). Male to female ratio was 6:10. After a single dose of ranibizumab injection, visual gain was noted in 14 of 17 study eyes. In two eyes, visual acuity was minimally reduced and no change was noted in the remaining eye with an initial visual acuity of hand motions. While pre-injection mean best-corrected visual acuity (BCVA) was 1.45 ±0.88 log Mar unit, post-injection mean BCVA was 1.00±0.68, 0.86 ±0.70, 0.80 ±0.71, 0.77 ±0.70, 0.77 ±0.70 log Mar unit respectively at the first week, first month, third month, sixth month and first year. In all patients, the mean RNFLT dramatically decreased after the injection during the follow- up. While pre-injection mean RNFLT was 210 ±38 µm, post-injection mean RNFLT was 162.11±40.2, 94±27, 71.23±22.5, 63 ±19 and 57 ±18 µm respectively at the first week, first month, third month, sixth month and first year. No injection related complication was noted during the follow-up period.
Intravitreal ranibizumab injection can be a treatment modality in eyes with acute NAION.
Intravitreal injection; nonarteritic anterior ischemic optic neuropathy; optic coherence tomography; optic nerve; ranibizumab.
Glioma risk has consistently been inversely associated with allergy history but not with smoking history despite putative biologic plausibility. Data from 855 high-grade glioma cases and 1,160 controls from 4 geographic regions of the United States during 1997–2008 were analyzed for interactions between allergy and smoking histories and inherited variants in 5 established glioma risk regions: 5p15.3 (TERT), 8q24.21 (CCDC26/MLZE), 9p21.3 (CDKN2B), 11q23.3 (PHLDB1/DDX6), and 20q13.3 (RTEL1). The inverse relation between allergy and glioma was stronger among those who did not (odds ratioallergy-glioma = 0.40, 95% confidence interval: 0.28, 0.58) versus those who did (odds ratioallergy-glioma = 0.76, 95% confidence interval: 0.59, 0.97; Pinteraction = 0.02) carry the 9p21.3 risk allele. However, the inverse association with allergy was stronger among those who carried (odds ratioallergy-glioma = 0.44, 95% confidence interval: 0.29, 0.68) versus those who did not carry (odds ratioallergy-glioma = 0.68, 95% confidence interval: 0.54, 0.86) the 20q13.3 glioma risk allele, but this interaction was not statistically significant (P = 0.14). No relation was observed between glioma risk and smoking (odds ratio = 0.92, 95% confidence interval: 0.77, 1.10; P = 0.37), and there were no interactions for glioma risk of smoking history with any of the risk alleles. The authors’ observations are consistent with a recent report that the inherited glioma risk variants in chromosome regions 9p21.3 and 20q13.3 may modify the inverse association of allergy and glioma.
allergy and immunology; glioma; hypersensitivity; polymorphism, single nucleotide
To compare the efficacy of low-fluence photodynamic therapy (PDT) combinations in the treatment of age-related macular degeneration (AMD).
Forty-five previously untreated eyes of 45 patients with exudative AMD whose best-corrected visual acuity (BCVA) was ≥0.3 (Snellen) were enrolled. 15 patients in Group I underwent low-fluence PDT (25J/cm2-300mW/cm2-83sec) and intravitreal pegaptanib combination, 15 patients in Group II underwent PDT (50J/cm2-600mW/cm2-83sec) and intravitreal pegaptanib combination while, 15 patients in Group III underwent intravitreal pegaptanib monotherapy. Complete ophthalmologic examinations were performed in pre and post treatment visits, and the results were statistically analised. A clinical activity score (CAS) was calculated by using changes in lesion size, amount of hemorrhage, staining pattern in FA and OCT measurement of intra/subretinal fluid. ≤ 3 logMAR lines of decrease in BCVA and decrease in CAS were considered as successful treatment.
The mean age of 19 female (42.2%) and 26 male (57.8%) patients was 72.82±8.02 years. Mean follow-up was 13.93±5.87 months. Lesion type was occult in 28 eyes (62.2%). Treatment success rates according to BCVA assessments were 86.7%, 80%, 60% and mean BCVA decrease were 0.3, 1.0, 2.2 logMAR lines in Group I, II and III, respectively (P>0.05). According to the changes in central macular thickness and CAS, no difference was found among the study groups (P=0.850 and P=0.811, respectively). Patients treated with combination regimens had lower intravitreal injection frequencies (P=0.015).
Combination regimen with intravitreal pegaptanib and low-fluence PDT seems to be safe and effective in stabilizing the clinical activity and BCVA in exudative AMD.
age-related macular degeneration; photodynamic therapy; low-fluence photodynamic therapy; anti-VEGF
To determine the prevalence of prediabetes and diabetes among rural and urban Malaysians.
RESEARCH DESIGN AND METHODS
This cross-sectional survey was conducted among 3,879 Malaysian adults (1,335 men and 2,544 women). All subjects underwent the 75-g oral glucose tolerance test (OGTT).
The overall prevalence of prediabetes was 22.1% (30.2% in men and 69.8% in women). Isolated impaired fasting glucose (IFG) and impaired glucose tolerance (IGT) were found in 3.4 and 16.1% of the study population, respectively, whereas 2.6% of the subjects had both IFG and IGT. Based on an OGTT, the prevalence of newly diagnosed type 2 diabetes was 12.6% (31.0% in men and 69.0% in women). The prediabetic subjects also had an increased prevalence of cardiovascular disease risk factors.
The large proportion of undiagnosed cases of prediabetes and diabetes reflects the lack of public awareness of the disease.
To examine the association between obesity and survival in patients with glioblastoma mutliforme (GBM)
Using a prospective design, 1,259 patients with previously untreated GBM were recruited between 1991 and 2008. Height and weight were self-reported or abstracted from medical records at study entry and used to calculate body mass index (BMI) [weight (kg)/[height (m)]2. Cox proportional models were used to estimate the risk of death associated with BMI as a continuous variable or categorized using established criteria (normal weight, 18.5–24.9 kg/m2; overweight, 25.0–29.9 kg/m2; obese, ≥30.0 kg/m2).
Median follow-up was 40 months, and 1,069 (85%) deaths were observed during this period. For all patients, minimal adjusted analyses indicated no significant association between BMI treated as a continuous variable and survival. Compared with patients with a BMI 18.5–24.9 kg/m2, the minimally adjusted HR for overall survival was 1.08 (95% CI, 0.94–1.24) for a BMI 25–29.9 kg/m2 and 1.08 (95% CI, 0.91–28) for a BMI ≥ 30.0 kg/m2. After additional adjustment for adjuvant therapy, the HR for those with a BMI of 25.0–29.9 kg/m2 was 1.14 (95% CI, 0.99–1.32) and 1.09 (95% CI, 0.91–1.30) for those with a BMI ≥30.0 kg/m2. No significant interactions were revealed for BMI and any demographic variables.
BMI was not associated with survival in newly diagnosed and previously untreated patients with GBM. Further research investigating the prognostic significance of alternative, quantitative measures of body habitus, and functional performance are required.
Epidemiology; Glioblastoma; Survival; Prognosis; Body weight; Energy balance
Hyperbaric oxygen (HBO2) exposure affects both oxidative and antioxidant systems. This effect is positively correlated with the exposure time and duration of the treatment. The present study aims enlightening the relation of HBO2 with oxidative/antioxidant systems when administered in a prolonged and repetitive manner in brain tissues of rats. Sixty rats were divided into 6 study (n = 8 for each) and 1 control (n = 12) group. Rats in the study groups were daily exposed 90-min HBO2 sessions at 2.8 ATA for 5, 10, 15, 20, 30 and 40 days. One day after the last session, animals were sacrificed; their whole brain tissue was harvested and dissected into three different regions as the outer grey matter (cortex), the inner white matter and cerebellum. Levels of lipid peroxidation and protein oxidation and activities of superoxide dismutase and glutathione peroxidase were measured in these tissues. Malondialdehyde, carbonylated protein and glutathione peroxidase levels were found to be insignificantly increased at different time-points in the cerebral cortex, inner white matter and cerebellum, respectively. These comparable results provide evidence for the safety of HBO treatments and/or successful adaptive mechanisms at least in the brain tissue of rats, even when administered for longer periods.
Allergies have been associated with decreased risk of glioma, but associations between duration and timing of allergies, and antihistamine use and glioma risk have been less consistent. The objective was to investigate this association by analyzing types, number, years since diagnosis, and age at diagnosis of allergies, and information on antihistamine usage, including type, duration, and frequency of exposure.
Self-report data on medically-diagnosed allergies and antihistamine use were obtained for 419 glioma cases and 612 hospital-based controls from Duke University and NorthShore University HealthSystem.
High- and low-grade glioma cases were statistically significantly less likely to report any allergy than controls (OR= 0.66, 95% CI: 0.49–0.87 and 0.44, 95% CI: 0.25–0.76, respectively). The number of types of allergies (seasonal, medication, pet, food, and other) was inversely associated with glioma risk in a dose-response manner (p-value for trend <0.05). Age at diagnosis and years since diagnosis of allergies were not associated with glioma risk. Oral antihistamine use was statistically significantly inversely associated with glioma risk, but when stratified by allergy status, remained significant only for those with high-grade glioma and no medically-diagnosed allergy.
All types of allergies appear to be protective with reduced risk for those with more types of allergies. Antihistamine use, other than in relationship with allergy status, may not influence glioma risk.
A comprehensive study of allergies and antihistamine use using standardized questions and biological markers will be essential to further delineate the biological mechanism that may be involved in brain tumor development.
Allergy; brain tumor; glioma; risk factor
EGFR and its constitutively activated variant EGFRvIII are linked to glioblastoma resistance to therapy, the mechanisms underlying this association, however, are still unclear. We report that in glioblastoma, EGFR/EGFRvIII paradoxically co-expresses with p53-upregulated modulator of apoptosis (PUMA), a proapoptotic member of the Bcl-2 family of proteins primarily located on the mitochondria. EGFR/EGFRvIII binds to PUMA constitutively and under apoptotic stress, and subsequently sequesters PUMA in the cytoplasm. The EGFR-PUMA interaction is independent of EGFR activation and is sustained under EGFR inhibition. A Bcl-2/Bcl-xL inhibitor that mimics PUMA activity sensitizes EGFR/EGFRvIII-expressing glioblastoma cells to Iressa. Collectively, we uncovered a novel kinase-independent function of EGFR/EGFRvIII that leads to tumor drug resistance.
EGFR; EGFRvIII; glioblastoma; apoptosis; PUMA; chemoresistance
A 21-year-old male with von Hippel-Lindau syndrome was treated for a retinal hemangioblastoma with photodynamic therapy (PDT). We employed the standard PDT protocol with doubled treatment duration. Seven days after the procedure, there were some perilesional intra- and subretinal hemorrhages and significant exudative retinal detachment extending inferiorly. Four weeks later, the hemorrhages had mostly cleared, and diminished subretinal fluid and partial regression of the hemangioblastoma were noted. PDT might induce temporary subretinal fluid accumulation in eyes with retinal hemangiomas.
Photodynamic Treatment; Retinal Capillary Hemangioblastoma; Retinal Detachment
Epidermal growth factor receptor (EGFR) plays an essential role in normal development, tumorigenesis and malignant biology of human cancers, and is known to undergo intracellular trafficking to subcellular organelles. Although several studies have shown that EGFR translocates into the mitochondria in cancer cells, it remains unclear whether mitochondrially localized EGFR has an impact on the cells and whether EGFRvIII, a constitutively activated variant of EGFR, undergoes mitochondrial transport similar to EGFR.
We report that both receptors translocate into the mitochondria of human glioblastoma and breast cancer cells, following treatments with the apoptosis inducers, staurosporine and anisomycin, and with an EGFR kinase inhibitor. Using mutant EGFR/EGFRvIII receptors engineered to undergo enriched intracellular trafficking into the mitochondria, we showed that glioblastoma cells expressing the mitochondrially enriched EGFRvIII were more resistant to staurosporine- and anisomycin-induced growth suppression and apoptosis and were highly resistant to EGFR kinase inhibitor-mediated growth inhibition.
These findings indicate that apoptosis inducers and EGFR-targeted inhibitors enhance mitochondrial translocalization of both EGFR and EGFRvIII and that mitochondrial accumulation of these receptors contributes to tumor drug resistance. The findings also provide evidence for a potential link between the mitochondrial EGFR pathway and apoptosis.
Emerging evidence indicate novel modes of EGFR signaling, notably, one involves EGFR nuclear translocalization and subsequent gene activation. To date, however, the significance of the nuclear EGFR pathway in glioblastoma (GBM) is unknown. Here, we report that EGFR and its constitutively activated variant EGFRvIII undergo nuclear translocalization in GBM cells, in which the former event requires EGF stimulation and the latter is constitutive. To gain insight into the impact of nuclear EGFR on gene expression in GBM, we created isogenic GBM cell lines, namely, U87MG-vector, U87MG-EGFR and U87MG-EGFRdNLS that, respectively, express the control vector, EGFR and nuclear-entry defective EGFR with a deletion of the nuclear localization signal (NLS). Microarray analysis shows that 19 genes, including, cyclooxygenase-2 (COX-2), to be activated in U87MG-EGFR cells but not in U87MG-EGFRdNLS and U87MG-vector cells. Subsequent validation studies indicate that COX-2 gene is expressed at higher levels in cells with EGFR and EGFRvIII than those with EGFRdNLS and EGFRvIIIdNLS. Nuclear EGFR and its transcriptional co-factor STAT3 associate with the COX-2 promoter. Increased expression of EGFR/EGFRvIII and activated STAT3 leads to synergistic activation of the COX-2 promoter. Promoter mutational analysis identified a proximal STAT3-binding site that is required for EGFR/EGFRvIII-STAT3 mediated COX-2 gene activation. In GBM tumors, an association exists between levels of COX-2, EGFR/EGFRvIII and activated STAT3. Together, these findings indicate the existence of the nuclear EGFR/EGFRvIII signaling pathway in GBM and its functional interaction with STAT3 to activate COX-2 gene expression, thus linking EGFR-STAT3 and EGFRvIII-STAT3 signaling axes to pro-inflammatory COX-2 mediated pathway.
EGFR; EGFRvIII; STAT3; COX-2; glioma; transcriptional regulation
Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor in adults. Current therapy includes surgery, radiation and chemotherapy with temozolomide (TMZ). Major determinants of clinical response to TMZ include methylation status of the O6-methylguanine-DNA methyltransferase (MGMT) promoter and mismatch repair (MMR) status. Though the MGMT promoter is methylated in 45% of cases, for the first nine months of follow-up, TMZ does not change survival outcome. Furthermore, MMR deficiency makes little contribution to clinical resistance, suggesting that there exist unrecognized mechanisms of resistance. We generated paired GBM cell lines whose resistance was attributed to neither MGMT nor MMR. We show that, responding to TMZ, these cells exhibit a decoupling of DNA damage response (DDR) from ongoing DNA damages. They display methylation-resistant synthesis in which ongoing DNA synthesis is not inhibited. They are also defective in the activation of the S and G2 phase checkpoint. DDR proteins ATM, Chk2, MDC1, NBS1 and gammaH2AX also fail to form discrete foci. These results demonstrate that failure of DDR may play an active role in chemoresistance to TMZ. DNA damages by TMZ are repaired by MMR proteins in a futile, reiterative process, which activates DDR signaling network that ultimately leads to the onset of cell death. GBM cells may survive genetic insults in the absence of DDR. We anticipate that our findings will lead to more studies that seek to further define the role of DDR in ultimately determining the fate of a tumor cell in response to TMZ and other DNA methylators.
glioblastomas multiforme; temozolomide; DNA damage response; resistance
The family of GLI zinc-finger transcription factors regulates the expression of genes involved in many important cellular processes, notably, embryonal development and cellular differentiation. The glioma-associated oncogene homolog 1 (GLI1) isoform, in particular, has attracted much attention because of its frequent activation in many human cancers and its interactions with other signaling pathways, such as, those mediated by K-RAS, TGF-β, EGFR and PKA. Here, we report the identification of a novel truncated GLI1 splice variant, tGLI1, with an in-frame deletion of 123 bases (41 codons) spanning the entire exon 3 and part of exon 4 of the GLI1 gene. Expression of tGLI1 is undetectable in normal cells but is high in glioblastoma multiforme (GBM) and other cancer cells. Although tGLI1 undergoes nuclear translocalization and transactivates GLI1-binding sites similar to GLI1, unlike GLI1, it is associated with increased motility and invasiveness of GBM cells. Using microarray analysis, we showed over 100 genes to be differentially expressed in tGLI1- compared to GLI1-expressing GBM cells, although both cell types expressed equal levels of known GLI1-regulated genes, such as, PTCH1. We further showed that one of the tGLI1 upregulated genes, CD24, an invasion-associated gene to be required for the migratory and invasive phenotype of GBM cells. These data provide conclusive evidence for a novel gain-of-function GLI1 splice variant that promotes migration and invasiveness of GBM cells and open up a new research paradigm on the role of the GLI1 pathway in malignancy.
Hedgehog pathway; GLI1; glioma; migration; invasion; CD24