Expression Of Glucose Transporter Protein-1 Research Articles

GLUT1 Promotes NLRP3 Inflammasome Activation of Airway Epithelium in Lipopolysaccharide-Induced Acute Lung Injury

Acute lung injury (ALI) is a devastating clinical disease caused by different factors, with high morbidity and mortality. It has been shown that lung injury and inflammation caused by lipopolysaccharide (LPS) can be modulated by NLRP3 inflammasome activation, yet its exact function within the airway epithelium is still unknown. Meanwhile, glucose transporter protein 1 (GLUT1) has been shown to contribute to a number of inflammatory illnesses, including ALI. The present study was aimed to assess GLUT1's function in NLRP3 inflammasome activation of airway epithelium in LPS-induced acute lung injury. BALB/c mice (5 mg/kg) and BEAS-2B cells (200 μg/mL) were respectively exposed to LPS, with or without GLUT1 antagonists (WZB117 or BAY876). LPS upregulated pulmonary expression of NLRP3 and GLUT1 in mice, which could be blocked by WZB117 or BAY876. Pharmacological inhibition of GLUT1 in vivo significantly attenuated lung tissue damage, neutrophil accumulation, and proinflammatory factors release (TNF-α, IL-6, and IL-1β) in LPS-exposed mice. Meanwhile, the activation markers of NLRP3 inflammasome (ASC, caspase-1, IL-1β, and IL-18) induced by LPS were also suppressed. In cultured BEAS-2B cells, LPS induced an increase in GLUT1 expression and triggers activation of the NLRP3 inflammasome, both of which were inhibited by GLUT1 antagonists. These results illustrated that GLUT1 participates in LPS-induced ALI, and promotes the activation of the NLRP3 inflammasome in airway epithelial cells.

ERK/PKM2 Is Mediated in the Warburg Effect and Cell Proliferation in Arsenic-Induced Human L-02 Hepatocytes.

This study aimed to investigate the potential role of pyruvate kinase M2 (PKM2) and extracellular regulated protein kinase (ERK) in arsenic-induced cell proliferation. L-02 cells were treated with 0.2 and 0.4 μmol/L As3+, glycolysis inhibitor (2-deoxy-D-glucose,2-DG), ERK inhibitor [1,4-diamino-2,3-dicyano-1,4-bis(2-aminophenylthio)-butadiene, U0126] or transfected with PKM2 plasmid. Cell viability, proliferation, lactate acid production, and glucose intake capacity were determined by CCK-8 assay, EdU assay, lactic acid kit and 2-deoxy-2-[(7-nitro-2,1,3-benzoxadiazol-4-yl) amino]-D-glucose (2-NBDG) uptake kit, respectively. Also, levels of PKM2, phospho-PKM2S37, glucose transporter protein 1 (GLUT1), lactate dehydrogenase A (LDHA), ERK, and phospho-ERK were detected using Western blot and the subcellular localization of PKM2 in L-02 cells was detected by immunocytochemistry (ICC). Treatment with 0.2 and 0.4 μmol/L As3+ for 48 h increased the viability and proliferation of L-02 cells, the proportion of 2-NBDG+ cell and lactic acid in the culture medium, and GLUT1, LDHA, PKM2, phospho-PKM2S37, and phospho-ERK levels and PKM2 in nucleus. Compared with the 0.2 μmol/L As3+ treatment group, the lactic acid in the culture medium, cell proliferation and cell viability, and the expression of GLUT1 and LDHA were reduced in the group co-treated with siRNA-PKM2 and arsenic or in the group co-treated with U0126. Moreover, the arsenic-increased phospho-PKM2S37/PKM2 was decreased by U0126. Therefore, ERK/PKM2 plays a key role in the Warburg effect and proliferation of L-02 cells induced by arsenic, and also might be involved in arsenic-induced upregulation of GLUT1 and LDHA. This study provides a theoretical basis for further elucidating the carcinogenic mechanism of arsenic.

GLUT-1 Expression in Breast Cancer.

Numerous studies have been conducted to predict the prognosis of breast cancers. The effect of glucose transporter protein 1 (GLUT-1), the main carrier protein responsible for glucose transport, was investigated in breast cancer patients. 170 patients operated for breast carcinoma were included in this study. We analysed the prognostic significance of GLUT-1 immune-expression in 149 patients without neoadjuvant therapy, and in 21 patients with neoadjuvant therapy. GLUT-1 expression was correlated with poor prognostic factors such as estrogen receptor and progesterone receptor negativity, high Ki-67 proliferation index, and high histological and nuclear grade (p < 0.001). GLUT-1 was expressed at a statistically higher rate in invasive ductal carcinomas, compared to invasive lobular carcinomas (p < 0.001), and was expressed at a higher rate in luminal B, human epidermal growth factor receptor 2 and triple-negative molecular subtypes compared to luminal A subtype tumors (p < 0.001). There was no statistically significant difference between GLUT-1 expression and presence of neoadjuvant therapy. Univariate survival analysis showed high GLUT1 expression was associated with low disease-free survival. GLUT-1 expression was found to be associated with poor pathological prognostic factors in breast carcinoma patients. The results suggest that GLUT-1 expression can be considered as a prognostic marker in breast cancers, and it may be used as a target molecule in personalized treatment approaches.

Down-Regulation of Hypoxia-Inducible Factor-1α and Downstream Glucose Transporter Protein-1 Gene by β-elemene Enhancing the Radiosensitivity of Lung Adenocarcinoma Transplanted Tumor.

PurposeTo study the effect of β-elemene on the radiosensitivity of A549 cell xenograft tumor and potential mechanisms by which β-elemene regulates the expression of hypoxia-inducible factor-1α (HIF-1α) and glucose transporter protein-1 (GLUT-1).MethodsUsing an A549 cell transplantation tumor model with male nude mice, we studied the effect of β-elemene on the radiosensitivity of non-small cell lung cancer (NSCLC). The expression of HIF-1α and GLUT-1 was detected by real-time PCR, Western blotting and immunohistochemistry. The relationship between the radiosensitivity of β-elemene and the expression of HIF-1α and GLUT-1 was analyzed.Resultsβ-elemene and radiotherapy intervened in the growth of transplanted tumors in varying degrees. The enhancement factor (EF=2.44>1) was calculated; β-elemene at 45 mg/kg had the most significant enhanced effect on radiosensitivity. When β-elemene was used in combination with radiation, the expression of HIF-1α and GLUT-1 was significantly decreased, and there was a positive correlation between the two genes.Conclusionβ-elemene exhibits a radiosensitizing effect on A549 cell xenograft tumor. The underlying molecular mechanism is probably associated with the down-regulation of HIF-1α and GLUT-1 expression, suggesting that β-elemene may directly or indirectly inhibit the expression of HIF-1α and GLUT-1. There is a positive significant correlation between expression of HIF-1α and GLUT-1. HIF-1α and downstream GLUT-1 could be used as a new target for the radiosensitization of NSCLC.

Relationships between expression of glucose transporter protein-1 and hypoxia inducible factor-1α, prognosis and 18F-FDG uptake in laryngeal and hypopharyngeal carcinomas.

BackgroundWe investigated the relationships between glucose transporter protein-1 (GLUT-1) and hypoxia inducible factor-1α (HIF-1α) expression, 2-deoxy-2-[18F]-fluoro-D-glucose (18F-FDG) uptake and prognosis in laryngeal and hypopharyngeal carcinomas patients.MethodsLevels of GLUT-1 and HIF-1α proteins were examined by immunohistochemistry in 55 patients with laryngeal and hypopharyngeal carcinomas. All patients underwent 18F-FDG positron emission tomography-computed tomography (PET/CT) before surgery.ResultsLevels of GLUT-1 and HIF-1α in laryngeal and hypopharyngeal carcinomas were significantly higher than in vocal cord polyps. GLUT-1 expression was correlated with HIF-1α expression (r=0.571, P<0.001) and SUVmax (r=0.495, P<0.001). HIF-1α expression was not correlated with SUVmax (P=0.199). HIF-1α expression was correlated with tumor size (r=0.351, P=0.009). Multivariate analysis demonstrated that treatment method (P=0.025), tumor size (P=0.008), GLUT-1 expression (P=0.032), and HIF-1α expression (P=0.032) were correlated with overall survival (OS).ConclusionsTreatment method, tumor size, GLUT-1 and HIF-1α levels were correlated with prognosis in laryngeal and hypopharyngeal carcinomas patients.

3-Bromopyruvate alleviates the development of monocrotaline-induced rat pulmonary arterial hypertension by decreasing aerobic glycolysis, inducing apoptosis, and suppressing inflammation.

Background:Pulmonary arterial hypertension (PH) is a progressive disease with limited therapeutic options, ultimately leading to right heart failure and death. Recent findings indicate the role of the Warburg effect (aerobic glycolysis) in the development of PH. However, the effect of the glycolysis inhibitor 3-bromopyruvate (3-BrPA) on the pathogenesis of PH has not been well investigated. This study aimed to determine whether 3-BrPA inhibits PH and its possible mechanism.Methods:PH was induced in adult Sprague-Dawley rats by a single intraperitoneal injection of monocrotaline (MCT). 3-BrPA, or phosphate-buffered saline (PBS) was administered via intraperitoneal injection every other day from the first day of MCT-injection to 4 weeks of follow-up, and indices such as right ventricular systolic pressure (RVSP), right ventricular hypertrophy index (RVHI), pulmonary arteriolar remodeling indicated by percent media thickness (% MT), lactate levels and glucose consumption, were evaluated. Pulmonary arteriolar remodeling and right ventricular hypertrophy were observed in hematoxylin-eosin-stained lung sections. Western blotting, immunohistochemistry, and/or immunofluorescence analyses were used to measure the expression of relevant proteins. A cytochrome C release apoptosis assay and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling staining were used to measure cell apoptosis.Results:MCT-induced PH showed a significant increase in glucose consumption (0 vs. 4 weeks: 0.87 ± 0.23 vs. 2.94 ± 0.47, P = 0.0042) and lactate production (0 vs. 4 weeks: 4.19 ± 0.34 vs. 8.06 ± 0.67, P = 0.0004). Treatment with 3-BrPA resulted in a concomitant reduction in glucose consumption (1.10 ± 0.35 vs. 3.25 ± 0.47, P = 0.0063), lactate production (5.09 ± 0.55 vs. 8.06 ± 0.67, P = 0.0065), MCT-induced increase in RVSP (39.70 ± 2.94 vs. 58.85 ± 2.32, P = 0.0004), pulmonary vascular remodeling (% MT, 43.45% ± 1.41% vs. 63.66% ± 1.78%, P < 0.0001), and right ventricular hypertrophy (RVHI, 38.57% ± 2.69% vs. 62.61% ± 1.57%, P < 0.0001) when compared with those of the PBS-treated group. 3-BrPA, a hexokinase 2 inhibitor, exerted its beneficial effect on PH by decreasing aerobic glycolysis and was also associated with inhibiting the expression of glucose transporter protein-1, inducing apoptosis, and suppressing inflammation.Conclusions:3-BrPA might have a potential beneficial effect on the PH treatment.

Evaluation of 18F-FDG PET/CT parameters for reflection of aggressiveness and prediction of prognosis in early-stage cervical cancer.

The aim of this study was to investigate the value of five typical fluorine-18-fluorodeoxyglucose PET/computed tomography (CT) parameters (including SUVmax, SUVmean, SUVpeak, MTV and TLG) in the reflecting aggressiveness and predicting prognosis in patients with early-stage cervical cancer. Our study enrolled 85 primary cervical cancer patients who underwent preoperative fluorine-18-fluorodeoxyglucose PET/CT scans. We investigated the association of five parameters derived from PET/CT with clinicopathological characteristics. Immunohistochemistry was utilized to evaluate the expression of glucose transporter protein-1 (GLUT-1), TP53-induced glycolysis and apoptosis regulator (TIGAR), and vascular endothelial growth factor (VEGF), and the correlation between PET/CT parameters and the expression of GLUT-1, TIGAR, and VEGF was analyzed. In addition, we also investigated the correlation between disease-free survival of cervical cancer patients and PET/CT parameters. The mean peak standardized uptake value (SUV)peak showed significant differences between all three International Federation of Gynecology and Obstetrics stages (P<0.001-0.032). The patients with lymph node metastasis had significantly higher SUVpeak, metabolic tumor volume (MTV), and total lesion glycolysis (TLG) compared with those with the absence of lymph node metastasis (P=0.038, 0.024, and 0.010, respectively). All the five parameters showed an association with tumor size and cervical stromal invasion depth. Immunohistochemistry results indicated that the SUVpeak showed the highest association with the expression of GLUT-1, TIGAR, and VEGF (rs=0.705, P<0.001, rs=-0.466, P<0.001, rs=0.580, P<0.001, respectively) in these five parameters. The survival analysis showed that SUVpeak, MTV, and TLG were correlated with disease-free survival (P=0.042, P<0.001, and P=0.004, respectively). Early-stage cervical cancer with high SUVpeak may reflect more aggressive behavior, and SUVpeak, MTV, and TLG can be used to predict the prognosis of early-stage cervical cancer patients. It may help with the choice of individualized treatment options.

The relationship between ¹⁸F-FDG uptake and oncogene in human laryngeal squamous cell carcinoma transplantation tumor animal model

Objective:The aim of this study is to explore the relationship between [F18] fluoro 2 deoxy d glucose ¹⁸F-FDG) uptake and oncogene expression in human laryngeal squamous cell carcinoma (LSCC) transplantation tumor.Method:Ten nude mice were randomly divided into two groups, 5 mice of each group. One group was injected subcutaneously with Hep-2 cells to establish xenograft model, and the other one was injected with HIF-RNAi-Hep-2 cells. All animals were conducted with positron emission tomography computed tomography PET scan. The average tumor T/N ratio was calculated. After PET scanning, tumor tissue was picked off for immunohistochemical examination.Result:The tumor imaging were both clear and there was no significant difference between these two groups (P> 0.05). The expression of hypoxia inducible factor 1 alpha (HIF-1α), glucose transporter protein-1 (GLUT-1) and vascular endothelial growth factor (VEGF) were high in Hep-2 group. The expression of HIF-1α was lower in HIF-RNAi-Hep-2 group (P< 0.05). But there was no significant difference about the expression of GLUT-1 and VEGF between these two groups (P> 0.05).Conclusion:The oncogenic pathway activated in LSCC is related with the uptake of ¹⁸F-FDG. The glycolysis gene and vascular factor maybe play a critical role.

Effect of gemcitabine on the uptake of 18F-fluorodeoxyglucose and 18F-fluorothymidine in lung adenocarcinoma A549 cells and the animal tumor model

Gemcitabine is the first-line drug for nonsmall cell lung cancer, and 18F-fluorodeoxyglucose. (18F-FDG) and 18F-fluorothymidine. (18F-FLT) are positron emission tomography. (PET) imaging agents. The aim of this study was to explore the effect of gemcitabine on the uptake of 18F-FDG and 18F-FLT in A549 cells and the animal tumor model. The inhibitory effects of gemcitabine on cell growth were determined by tetrazolium blue method, and uptake rates of 18F-FDG and 18F-FLT were determined under the same conditions. The adenocarcinoma-bearing nude mice before and after gemcitabine treatments were performed microPET imaging with 18F-FDG and 18F-FLT. Hematoxylin and eosin staining and immunohistochemical analysis of tumor specimens were conducted. After the administration of gemcitabine, positive correlations were observed between inhibition of 18F-FDG or 18F.FLT uptake and cell growth. (r = 0.957 or 0.981, P < 0.01). SUVmax values by 18F-FDG in the tumor, before and after administration of gemcitabine at the dose of 60 mmol/L, revealed an increase by. (35.83 ± 10.58) %. After administration of 120 mmol/L gemcitabine, the SUVmax values decreased by (12.37 ± 7.33) %. The SUVmax values by 18F-FLT at the dose of 60 mmol/L gemcitabine revealed a decrease by (56.47 ± 10.83) %. Pathological staining showed obvious vasodilation and invasion of lymphocytes and plasma cells at the dose of 60 mmol/L, and the expression of glucose transporter protein-1, Ki-67 and proliferating cell nuclear antigen in tumor cells were inhibited. 18F-FLT imaging can assess the proliferation of tumor cells and 18F-FDG imaging can reflect the changes of the tumor microenvironment after administration of gemcitabine.

Expression of glucose transporter protein 1 and p63 in serous ovarian tumor.

It has been shown that glycolytic metabolism is increased in malignant cells. Cancer cell growth is an energy-related process supported by an increased glucose metabolism. In addition, p63, a known homolog of p53, is expressed predominantly in basal cell and squamous cell carcinomas. The purpose of this study was to evaluate the expression of glucose transporter protein 1 (GLUT1) and p63 in patients with serous ovarian tumor (benign, borderline and malignant) and study their close relationship with the malignant transformation of serous ovarian tumors. Two hundred formalin-fixed, paraffin-embedded sections were immunostained with rabbit anti-GLUT1 polyclonal antibody and mouse anti-p63 monoclonal antibody using the streptavidin-biotin method. The samples were as follows: 40 normal ovarian tissues, 40 serous cystadenomas, 40 borderline serous cystadenomas and 80 serous cystadenocarcinomas were stained. Normal ovarian tissues showed completely negative staining for GLUT1 and p63. However, from benign serious cystadenomas, borderline cystadenomas to cystadenocarcinomas, the expression of GLUT1 and p63 grew stronger (P < 0.05). Moreover, the intensity staining of GLUT1 maintained a significant association with the expression of p63 (P < 0.05). In χ²-test analysis, expression of borderline cystadenomas and cystadenocarcinomas, intraperitoneal implants, ascites, lymph node status and International Federation of Gynecology and Obstetrics (FIGO) stage and GLUT1 expression levels have an appalling significance (P < 0.05), while FIGO stage, intraperitoneal implants and lymph node status except patient age and ascites have a statistical significance with the expression of p63 levels (P < 0.05). Our findings show a progressive increase in the expression of GLUT1 and p63 from the benign serous cystadenomas, borderline cystadenomas to cystadenocarcinomas. Overexpression of GLUT1 and p63 are associated with the histology FIGO stage and metastasis of the tumors. These data suggested that the expression of GLUT1 and p63 may be closely related to the malignant transformation of serous ovarian tumors. However, the relative importance of GLUT1 and p63 in ovarian serous tumor development and tumorigenesis remains mostly unclear and awaits further investigation.

Protective role of Vitamin D in diabetic rats

Objective To investigate the protective effect of Vitamin D (VitD) on diabetic rats, and whether the protective mechanism is associated with the expression of nuclear factor kappa B (NF-κB) and insulin resistance (IR). Methods Diabetic Wistar rats were established by intraperitoneal injection of streptozotocin, and randomly divided into diabetic group, VitD treatment group(treatment group). Normal rats were served as normal control group.Treatment group was treated with VitD for 8 weeks.The levels of 24 h urinary protein (24 h UP), fasting plasma glucose (FPG), plasma insulin (p-Ins), plasma adiponectin (p-Adi), plasma glucagon (p-Gln) were measured.Tumor necrosis factor alpha (TNF-α), monocyte chemotactic protein 1 (MCP-1), interleukin-6 (IL-6) were determinated in renal cortical homogenate.The activity of NF-κB was evaluated by electrophoretic mobility shift assay.The mRNA expressions of glucose transporter protein 1 (GLUT1) and GLUT4 in renal cortex were detected by reverse transcriptase(RT)-PCR.Western blot analysis was performed to measure the phosphorylation of NF-κB and its inhibitor I kappa B-alpha (IκBα), insulin receptor substrate protein 1 (IRS1), phosphatidylinositol 3 kinase (PI3K), p38 mitogen activated protein kinase (p38MAPK). Results Compared with the normal control group, 24 h UP, FPG, p-Ins, p-Gln were significantly increased, and inflammatory markers and the expression of GLUT1 elevated in renal cortex in DM group, there were significant differences(all P<0.01). The activity of NF-κB (P<0.01) and the phosphorylation of NF-κB p65 and p38MAPK were elevated (all P<0.01), and phosphorylation of IκBα, IRS1, PI3K were decreased (all P<0.05, 0.01) in diabetic group compared with those of normal control group.VitD treatment could ameliorate urine protein, increase p-Adi, reduce inflammatory markers and NF-κB activity (P<0.01), maintain GLUT1 expression, but had no effect on GLUT4 expression in renal cortical, attenuate NF-κB p65, p38MAPK phosphorylation (all P<0.05), partly restore IκBα, IRS1, PI3K phosphorylation in diabetic rats (all P<0.05, 0.01). Conclusions Protective role of VitD is associated with inhibiting the expression of NF-κB and reducing the insulin resistance in diabetes. Key words: Vitamin D; Diabetes; Nuclear factor kappa B; Insulin resistance

Silencing of GLUT-I inhibits sensitization of oral cancer cells to cisplatin during hypoxia

Background During tumor development, cells are exposed to a hypoxic microenvironment. Tumor hypoxia also has a profound influence on the sensitivity of cancer chemotherapy. The objective of this study was to investigate the mechanism of cisplatin (CDDP) resistance of oral squamous cell carcinoma (OSCC) cells under hypoxia by analyzing gene expression profiles to identify key genes and factors involved. Methods Cell viability was measured following culture of the cells in the presence or absence of CDDP, under normoxic or hypoxic conditions, using a CCK-8 assay. Analysis of the expression of HIF target genes in hypoxia-treated cells was performed using an HIF-regulated cDNA plate array. Changes in the mRNA expression of selected HIF target genes were analyzed using RT-PCR, and changes in the protein levels of these genes were analyzed by Western blotting. Tumor cell apoptosis was assessed by flow cytometry. Results The OSCC cell lines responded differently to CDDP under normoxic and hypoxic conditions. The expression of glucose transporter protein-1 (GLUT-1) was up-regulated in human squamous cell carcinoma of mouth (HSC-2) cells under hypoxia. Furthermore, there was little correlation between the cisplatin sensitivity of human squamous cell carcinoma of tongue (SAS) in normoxia and hypoxia. After GLUT-1 knockdown, CDDP treatment resulted in increased rates of apoptosis under hypoxia as compared with normoxia in cell lines HSC-2, Ca9-22, and SAS (P = 0.025). Conclusion The results of this study suggest that knockdown of GLUT-1 inhibits sensitization of oral squamous cells to CDDP during hypoxia in HSC-2, Ca9-22, and SAS cells.

Silencing of GLUT‐1 inhibits sensitization of oral cancer cells to cisplatin during hypoxia

During tumor development, cells are exposed to a hypoxic microenvironment. Tumor hypoxia also has a profound influence on the sensitivity of cancer chemotherapy. The objective of this study was to investigate the mechanism of cisplatin (CDDP) resistance of oral squamous cell carcinoma (OSCC) cells under hypoxia by analyzing gene expression profiles to identify key genes and factors involved. Cell viability was measured following culture of the cells in the presence or absence of CDDP, under normoxic or hypoxic conditions, using a CCK-8 assay. Analysis of the expression of HIF target genes in hypoxia-treated cells was performed using an HIF-regulated cDNA plate array. Changes in the mRNA expression of selected HIF target genes were analyzed using RT-PCR, and changes in the protein levels of these genes were analyzed by Western blotting. Tumor cell apoptosis was assessed by flow cytometry. The OSCC cell lines responded differently to CDDP under normoxic and hypoxic conditions. The expression of glucose transporter protein-1 (GLUT-1) was up-regulated in human squamous cell carcinoma of mouth (HSC-2) cells under hypoxia. Furthermore, there was little correlation between the cisplatin sensitivity of human squamous cell carcinoma of tongue (SAS) in normoxia and hypoxia. After GLUT-1 knockdown, CDDP treatment resulted in increased rates of apoptosis under hypoxia as compared with normoxia in cell lines HSC-2, Ca9-22, and SAS (P = 0.025). The results of this study suggest that knockdown of GLUT-1 inhibits sensitization of oral squamous cells to CDDP during hypoxia in HSC-2, Ca9-22, and SAS cells.

Association of tumor hypoxia with lower survival after radiotherapy for muscle-invasive bladder cancer.

292 Background: Hypoxia may influence clinical outcomes in many human tumors, including muscle invasive urothelial carcinoma (UC) of the bladder. Hypoxia inducible factor-1 (HIF1), glucose transporter protein-1 (GLUT1) and carbonic anhydrase IX (CAIX) are all upregulated by hypoxia. This study examined the relationship between biomarkers of hypoxia or proliferation (Ki67) and clinical outcome in UC patients treated with radiotherapy (RT). Methods: 70 patients were treated with curative intent using RT alone or RT plus neoadjuvant or concurrent chemotherapy (RTCT) between 1997 and 2006. Archived, paraffin-embedded, transurethral biopsy material was available for 44 patients (35 men and 9 women). Tumor was clinically confined to the bladder wall (cT2) in 28 patients and involved perivesical tissues or adjacent organs (cT3/T4) in 16. Tissue micro-arrays were constructed and immunohistochemical analyses performed to assess biomarker expression, which was scored independently by 3 observers in a semi-quantitative manner. The biomarker scores were correlated with clinico-pathologic data extracted from the medical record and with patient outcome. Median follow-up was 4.2 years. Results: HIF1, GLUT1 and CAIX expression was seen in 43%, 96% and 65% of tumors respectively. The median Ki67 expression was 60%. There were no correlations between biomarkers expression and clinico-pathologic prognostic factors. The cystoscopic complete response rate was 71% at 2-3 months post-RT. The 4-year actuarial local control (LC) and overall survival (OS) rates were 48% and 55% respectively. High Ki67 expression was strongly predictive of better LC by univariate and multivariate analyses (70% vs 20% at 4 years, HR 0.95, p= 0.002), whereas no hypoxic marker predicted LC. High CAIX expression was predictive of inferior OS by multivariate analysis (HR 1.6, p=0.04). Conclusions: Tumor hypoxia influences the outcome of patients with bladder cancer treated with RT or RTCT and is a potential therapeutic target. High Ki67 has been reported previously to be a marker of sustained LC in the bladder following RTCT but the biologic mechanism underlying this association is unknown and requires further investigation.

Congenital Transmission by Protozoan

Congenital transmission by protozoan parasites is a worldwide important public health problem. Congenital infections affects the mother and the fetus or newborn. It is still surprising that despite the abundant immunoepidemiological knowledge of congenital transmission of protozoan parasite, no definite etiology or predictive diagnostic tests have been identified. Understanding the mechanisms by which host/parasites infection and interaction occurs is one of the most important topics that will help find specific biomarkers of infection, prevent congenital transmission, maintain the health of the newborn, and develop safe and efficient treatments. In addition, understanding of the biological mechanisms of host/parasite interactions will facilitate protection of mothers and their families and reduce costs in health services. Moreover, this will lead to gain insight into epidemiological aspects and association with other pathologies and preserve the wellbeing of the newborn. In this special issue we have invited a few papers that address such issues. Congenital malaria is underestimated and usually associated with low-density cord parasitemia. However, it is increasingly recognized as a potentially serious complication of maternal malaria during pregnancy in Sub-Saharian Africa. Earliest epidemiological studies have reported prevalence varying widely in malaria-endemic areas from 0% to 33%. At birth, infections are usually asymptomatic with low parasitemia and the diagnosis by microscopy is often missed. Infection may occur by transplacental passage of parasites during disruption of the placental barrier at the time of delivery, with subsequent clinical illness in the newborn baby. A paper of this issue assessed the prevalence of congenital malaria during the dry season (period of low-mosquito density and low-malaria incidence) using peripheral and cord blood smears of new born babies in association with peripheral and placental blood smears of, respectively, near-term and term pregnant women. This study is interesting because the authors found that congenital malaria is not rare in their study area. Another paper is a literature review on the prevalence, burden, diagnosis, prevention, and control of congenital malaria in Sub-Saharian Africa. This review is of interest to individuals working in clinics and laboratory diagnostic of malaria but also to national governments and partners in Sub-Saharian Africa. The authors highlight the challenges in parasitological and clinical diagnosis, the challenges in prevention with the use of intermittent preventive treatment (IPT) and insecticide-treated nets (ITNs) and provides recommendation on how to strengthen the health system in Africa. Another paper reports the prevalence of transplacental malaria in Burkina Faso, a West African country, and determines the real burden of transplacental transmission. The authors show associations between levels of parasite in the maternal, placental, and umbilical cord blood. All papers about malaria published in this special issue show the interest of pursuing investigations for a better understanding of vertical transmission by malarial parasites. During congenital Chagas transmission, the parasite reaches the fetus by crossing the placental barrier. In the past few years congenital transmission of T. cruzi has increasingly become more important, and partly responsible for the “globalization of Chagas' disease,” constituting a public health problem of increasing relevance. The fact that only a percentage of the infected mothers transmit parasites to their fetuses raises the question of the ability of the placenta as well as the immunological status of mother and fetus/newborn to impair the parasite transmission. Therefore, it is thought that congenital Chagas disease is the product of a complex interaction between the parasite, the maternal, and fetus/newborn immune responses, and placental factors. Additionally, a paper of this special issue constitutes a morphological analysis by immunohistochemical and histochemical methods of placentas from women with chronic Chagas' disease. T. cruzi-infected placentas present destruction of the syncytiotrophoblast and villous stroma, selective disorganization of the basal lamina, and disorganization of collagen I in villous stroma. Changes in the extracellular matrix of placental tissues, together with the immunological status of mother and fetus, and parasite load may determine the probability of congenital transmission of T. cruzi. Another paper assayed the effect of T. cruzi on glucose transporter protein-1 (GLUT1), which is the main isoform involved in transplacental glucose transport. High glucose media as well as T. cruzi infection reduce GLUT1 expression. The effect of T. cruzi infection on GLUT1 expression may explain some of the clinical manifestation of congenital Chagas disease. Finally, a paper analyzing the congenital transmission of Chagas disease is a review about the role of possible local placental factors that contribute to the vertical transmission of the parasite. Additionally, in that review different available methods for studying the congenital transmission of T. cruzi are analyzed. In that context, the ex vivo infection of human placental chorionic villi by T. cruzi trypomastigotes constitutes an excellent tool for studying parasite infection strategies as well as possible local antiparasitic mechanisms. Ricardo E. Fretes Ulrike Kemmerling Demba Sarr

In VitroInfection ofTrypanosoma cruziCauses Decrease in Glucose Transporter Protein-1 (GLUT1) Expression in Explants of Human Placental Villi Cultured under Normal and High Glucose Concentrations

Trypanosoma cruzi, the etiologic Chagas' disease agent, induces changes in protein pattern of the human placenta syncytiotrophoblast. The glucose transporter protein-1 (GLUT1) is the primary isoform involved in transplacental glucose transport. We carried out in vitro assays to determine if T. cruzi infection would induce changes in placental GLUT1 protein expression under normal and high concentration of glucose. Using Western blot and immunohistological techniques, GLUT1 expression was determined in normal placental villi cultured under normal or high concentrations of glucose, with or without in vitro T. cruzi infection, for 24 and 48 hours. High glucose media or T. cruzi infection alone reduced GLUT1 expression. A yet more accentuated reduction was observed when infection and high glucose condition took place together. We inform, for the first time, that T. cruzi infection may induce reduction of GLUT1 expression under normal and high glucose concentrations, and this effect is synergic to high glucose concentrations.

Impairment of brain endothelial glucose transporter by methamphetamine causes blood-brain barrier dysfunction

BackgroundMethamphetamine (METH), an addictive psycho-stimulant drug with euphoric effect is known to cause neurotoxicity due to oxidative stress, dopamine accumulation and glial cell activation. Here we hypothesized that METH-induced interference of glucose uptake and transport at the endothelium can disrupt the energy requirement of the blood-brain barrier (BBB) function and integrity. We undertake this study because there is no report of METH effects on glucose uptake and transport across the blood-brain barrier (BBB) to date.ResultsIn this study, we demonstrate that METH-induced disruption of glucose uptake by endothelium lead to BBB dysfunction. Our data indicate that a low concentration of METH (20 μM) increased the expression of glucose transporter protein-1 (GLUT1) in primary human brain endothelial cell (hBEC, main component of BBB) without affecting the glucose uptake. A high concentration of 200 μM of METH decreased both the glucose uptake and GLUT1 protein levels in hBEC culture. Transcription process appeared to regulate the changes in METH-induced GLUT1 expression. METH-induced decrease in GLUT1 protein level was associated with reduction in BBB tight junction protein occludin and zonula occludens-1. Functional assessment of the trans-endothelial electrical resistance of the cell monolayers and permeability of dye tracers in animal model validated the pharmacokinetics and molecular findings that inhibition of glucose uptake by GLUT1 inhibitor cytochalasin B (CB) aggravated the METH-induced disruption of the BBB integrity. Application of acetyl-L-carnitine suppressed the effects of METH on glucose uptake and BBB function.ConclusionOur findings suggest that impairment of GLUT1 at the brain endothelium by METH may contribute to energy-associated disruption of tight junction assembly and loss of BBB integrity.

Significance of GLUT1 expression in adenocarcinoma and adenoma of the ampulla of Vater

The expression of glucose transporter protein 1 (GLUT1) in malignant tumors is increased due to the higher metabolic needs of the proliferating cell populations. Aberrant GLUT1 expression is exhibited in a wide spectrum of epithelial malignancies and their precursors, which occur with low frequency and intensity; aberrant GLUT1 expression does not occur in normal epithelial cells. The expression of GLUT1 in tumors of the ampulla of Vater was evaluated on immunohistochemistry, and the relationships of GLUT1 expression to histological parameters and p53 expression were analyzed. Twenty-one (58.3%) of 36 adenocarcinomas and three (17.6%) of 17 adenomas had GLUT1 immunoreactivity. None of the regenerating or normal epithelia had any immunoreactivity. No significant relationships were found between GLUT1 expression and histological parameters or p53 expression. It was found that histological subtypes originated from different epithelium were strongly related to different macroscopic types. In the ampulla of Vater, GLUT1 expression was associated with malignant change, and might be a useful marker of malignancy.

Expression of Glucose Transporter Protein-1 (Glut-1) in Ocular Surface Squamous Neoplasia

To examine the expression of glucose transporter protein-1 (GLUT-1) in ocular surface squamous neoplasia and to study its relationship with degree of neoplasia and cell proliferation index (Ki-67 labeling index). Twelve cases diagnosed as ocular surface squamous neoplasia (4 invasive and 8 intraepithelial tumors) at Inonu University Faculty of Medicine, Department of Pathology, were included in this study. There were 3 squamous cell carcinomas, 1 basosquamous cell carcinoma, and 8 conjunctival intraepithelial neoplasms. Immunohistochemically, GLUT-1 and Ki-67 antibody staining were performed. GLUT-1 membranous immunoreactivity was seen in all tumors except in 1 case. GLUT-1 immunostaining was observed in all layers of the neoplastic epithelium of squamous cell carcinoma. Intense staining for GLUT-1 was determined in the upper two thirds of the severe dysplastic squamous epithelium. Although immunoreactivity for Ki-67 nuclear antigen was present throughout the epithelium, it was higher in the lower two thirds. Ki-67 labeling index ranged between 6% and 80%, and the mean value was 35% for invasive tumors and 20% for intraepithelial tumors. Marked GLUT-1 and Ki-67 immunoreactive cells throughout the neoplastic epithelium of ocular surface squamous neoplasia were observed. In most cases, it was observed that GLUT-1 expression was severe in cases having >10% Ki-67 labeling index. These findings indicate that glucose uptake was increased in dysplastic cells, especially by GLUT-1. To our knowledge, this is the first study on the subject in the literature, and further studies with more cases are needed with GLUT-1 and other GLUT members.

GLUT-1-independent infection of the glioblastoma/astroglioma U87 cells by the human T cell leukemia virus type 1

The human glucose transporter protein 1 (GLUT-1) functions as a receptor for human T cell leukemia virus (HTLV). GLUT-1 is a twelve-transmembrane cell surface receptor with six extracellular (ECL) and seven intracellular domains. To analyze HTLV-1 cytotropism, we utilized polyclonal antibodies to a synthetic peptide corresponding to the large extracellular domain of GLUT-1. The antibodies caused significant blocking of envelope (Env)-mediated fusion and pseudotyped virus infection of HeLa cells but had no significant effect on infection of U87 cells. This differential effect correlated with the detection of high-level surface expression of GLUT-1 on HeLa cells and very weak staining of U87 cells. To investigate this in terms of viral cytotropism, we cloned GLUT-1 cDNA from U87 cells and isolated two different versions of cDNA clones: the wild-type sequence (encoding 492 residues) and a mutant cDNA with a 5-base pair deletion (GLUT-1Δ5) between nucleotides 1329 and 1333. The deletion, also detected in genomic DNA, resulted in a frame-shift and premature termination producing a truncated protein of 463 residues. Transfection of the wild-type GLUT-1 but not GLUT-1Δ5 cDNA into CHO cells resulted in efficient surface expression of the human GLUT-1. Co-expression of GLUT-1 with GLUT-1Δ5 produces a trans-inhibition by GLUT-1Δ5 of GLUT-1-mediated HTLV-1 envelope (Env)-mediated fusion. Co-immunoprecipitation experiments demonstrated physical interaction of the wild-type and mutant proteins. Northern blot and RT-PCR analyses demonstrated lower GLUT-1 RNA expression in U87 cells. We propose two mechanisms to account for the impaired cell surface expression of GLUT-1 on U87 cells: low GLUT-1 RNA expression and the formation of GLUT-1/GLUT-1Δ5 heterodimers that are retained intracellularly. Significant RNAi-mediated reduction of endogenous GLUT-1 expression impaired HTLV-1 Env-mediated fusion with HeLa cells but not with U87 cells. We propose a GLUT-1-independent mechanism of HTLV-1 infection of U87 cells. The results may have important implications for HTLV-1 neurotropism and pathogenesis.