Translational Research

2012-05-01

Role of advanced glycation endproducts and glyoxalase I in diabetic peripheral sensory neuropathy

Megan Jack, Douglas Wright — 2012-01-11

Diabetic neuropathy is the most common and debilitating complication of diabetes mellitus with more than half of all patients developing altered sensation as a result of damage to peripheral sensory neurons. Hyperglycemia results in altered nerve conduction velocities, loss of epidermal innervation, and development of painful or painless signs and symptoms in the feet and hands. Current research has been unable to determine whether a patient will develop insensate or painful neuropathy or be protected from peripheral nerve damage all together. One mechanism that has been recognized to have a role in the pathogenesis of sensory neuron damage is the process of reactive dicarbonyls forming advanced glycation endproducts (AGEs) as a direct result of hyperglycemia. The glyoxalase system, composed of the enzymes glyoxalase I (GLO1) and glyoxalase II, is the main detoxification pathway involved in breaking down toxic reactive dicarbonyls before producing carbonyl stress and forming AGEs on proteins, lipids, or nucleic acids. This review discusses AGEs, GLO1, their role in diabetic neuropathy, and potential therapeutic targets of the AGE pathway.

Molecular profiling of individual tumor cells by hyperspectral microscopic imaging

2011-09-05

We developed a hyperspectral microscopic imaging (HMI) platform that can precisely identify and quantify 10 molecular markers in individual cancer cells in a single pass. The exploitation of an improved separation of circulating tumor cells and the application of HMI provided an opportunity (1) to identify molecular changes in these cells, (2) to recognize the coexpression of these markers, (3) to pose an important opportunity for noninvasive diagnosis, and (4) to use targeted therapy. We balanced the intensity of 10 fluorochromes bound to 10 different antibodies, each specific to a particular tumor marker, so that the intensity of each fluorochrome can be discerned from overlapping emissions. Using 2 touch preps from each primary breast cancer, the average molecular marker intensities of 25 tumor cells gave a representative molecular signature for the tumor despite some cellular heterogeneity. The intensities determined by the HMI correlate well with the conventional 0–3+ analysis by experts in cellular pathology. Because additional multiplexes can be developed using the same fluorochromes but different antibodies, this analysis allows quantification of many molecular markers on a population of tumor cells. HMI can be automated completely, and eventually, it could allow the standardization of protein biomarkers and improve reproducibility among clinical pathology laboratories.

Genetic analysis of the promoter region of the GATA4 gene in patients with ventricular septal defects

Guanghua Wu, Jiping Shan, Shuchao Pang, Xiaodan Wei, Hao Zhang, Bo Yan — 2011-11-24

Ventricular septal defects (VSDs) are the most common type of congenital heart diseases (CHDs). To date, the genetic causes for sporadic VSDs remain largely unknown. GATA transcription factor 4 (GATA4) is a zinc-finger transcription factor that is expressed in developing heart and adult cardiomyocytes. Mutations in the coding region of the GATA4 gene have been identified in CHD patients, including VSD. As the GATA4 factor is a dosage-sensitive regulator, we hypothesized that the promoter region variants of the GATA4 gene may be genetic causes of VSD. In this study, we analyzed the promoter region of the GATA4 gene by bidirectional sequencing in 172 VSD patients and 171 healthy controls. The results showed that 5 heterozygous sequence variants (NG_008177:g.4071T>C, NG_008177:g.4148C>A, NG_008177:g.4566C>T, NG_008177:g.4653G>T, and NG_008177:g.4690G>deletion) within the promoter region of the GATA gene were identified in 5 VSD patients, but in none of controls. One heterozygous sequence variant (g.4762C>A) was found only in one control, which may have no functional significance. A functional analysis revealed that the transcriptional activity of variant NG_008177:g.4566C>T was reduced significantly, whereas the transcriptional activities of the variants (NG_008177:g.4071T>C, NG_008177:g.4148C>A, NG_008177:g.4653G>T, and NG_008177:g.4690G>deletion) were increased significantly compared with the wild-type GATA4 gene promoter. As GATA4 is a dosage-sensitive regulator during development, our data suggest that these sequence variants within the promoter region of the GATA4 gene may contribute to the VSD etiology by altering its gene expression. Additional studies in experimental animals will deepen our understanding of the genetic basis of VSD and shed light on designing novel molecular therapies for adult VSD patients carrying these variants.

Uncoupling protein-2 expression and effects on mitochondrial membrane potential and oxidant stress in heart tissue

2011-12-05

Myocardial uncoupling protein (UCP)-2 is increased with chronic peroxisome proliferator-activated receptor γ (PPARγ) stimulation, but the effect on membrane potential and superoxide is unclear. Wild-type (WT) and UCP-2 knockout (KO) mice were given a 3-week diet of control (C) or the PPARγ agonist pioglitazone (PIO; 50 μg/g-chow per day). In isolated mitochondria, UCP-2 content by Western blots, membrane potential (ΔΨm) by tetraphenylphosphonium (TPP), and relative superoxide levels by dihydroethidium (DHE) were measured. Oxygen respiration was determined at baseline and after 10 min anoxia-reoxygenation. PIO induced a 2-fold increase in UCP-2 and nuclear-bound PGC1α in WT mice with no UCP-2 expression in KO mice. Mitochondrial ΔΨm from WT mice on C and PIO diets was –166 ± 4 mV and –147 ± 6 mV, respectively (P < 0.05). These values were lower than in UCP-2 KO mice on C and PIO (–180 ± 4 mV and –180 ± 4 mV, respectively; P < 0.05). Maximal complex III inhibitable superoxide from WT mice on C and PIO diets was 22.5 ± 1.3 and 17.8 ± 1.1 AU, respectively (P < 0.05), and were lower than UCP-2 KO on C and PIO (32.9 ± 2.3 and 29.2 ± 1.9 AU, respectively; P < 0.05). Postanoxia, the respiratory control index (RCI) in mitochondria from WT mice with and without PIO was 2.5 ± 0.3 and 2.4 ± 0.2, respectively, and exceeded that of UCP-2 KO mice on C and PIO (1.2 ± 0.1 and 1.4 ± 0.1, respectively; P < 0.05). In summary, chronic PPARγ stimulation leads to depolarization of the inner membrane and reduced superoxide of isolated heart mitochondria, which was critically dependent on increased expression of UCP-2. Thus, UCP-2 expression affords resistance to brief anoxia-reoxygenation.

Chronic kidney disease as a risk factor for acute coronary syndromes in patients presenting to the emergency room with chest pain

2012-01-10

We sought to determine whether persons with intermediate risk factors for cardiovascular disease presenting to an emergency department with chest pain and chronic kidney disease (CKD) were triaged effectively by chest pain units (CPUs). CPUs evaluate patients with intermediate risk and acute chest pain effectively. CKD is a risk factor for poor outcomes once cardiovascular disease has developed. However, current algorithms to risk stratify patients with acute chest pain do not include renal function. A total of 408 patients enrolled previously in the CHEER study of intermediate risk patients with chest pain, assigned randomly to hospitalization or observation in a CPU where an estimated glomerular filtration rate (GFR) was available, were included. No difference was found in short-term outcomes of patients including in-hospital death, myocardial infarction, or coronary revascularization based on renal function. For the 205 patients randomized to the CPU, the rate of admission to the hospital was significantly higher in the group with CKD compared with the group with normal renal function (68.2 vs 48.2%, P = 0.007). In a multivariate analysis, decreased renal function was not associated with adverse short-term outcomes. On 5 years follow-up, the overall long-term mortality was significantly higher in the group with CKD (14.1% vs 5.5%, P = 0.003). We concluded that CKD is a strong predictor of hospitalization and overall long-term mortality in patients presenting with chest pain to the emergency department. Current risk factor stratification scoring systems should consider CKD as a predictor of increased risk in patients with chest pain.

Genetic and immune predictors for hypersensitivity syndrome to antiepileptic drugs

Manuela G. Neuman, Lawrence Cohen, Radu M. Nanau, Paul A. Hwang — 2012-01-27

Hypersensitivity syndrome reactions (HSR) to antiepileptic drugs (AED) are associated with severe clinical cutaneous adverse reactions (SCAR). We aimed (1) to assess HSRs to AEDs using the in vitro lymphocyte toxicity assay (LTA) in patients who manifested HSRs clinically; (2) to correlate LTA results with the clinical syndrome; (3) to correlate LTA results with the human leukocyte antigen (HLA) allele B∗1502 (HLA-B∗1502) positivity in a Han Chinese-Canadian population; and (4) to determine the cytokine network in this population. Patients that developed fever and cutaneous eruptions in the presence or absence of organ involvement within 8 weeks of exposure to carbamazepine (CBZ), phenytoin (PHY), or lamotrigine (LTG) were enrolled. Control patients received AEDs without presenting HSR. We investigated 10 CBZ-HSR patients (4 with Stevens-Johnson syndrome [SJS]), 24 CBZ-controls, 10 PHY-HSR patients (4 with drug-induced liver injury [DILI]), 24 PHY-controls,6 LTG-HSR patients (1 with SJS and 1 with DILI), and 24 LTG-controls. There were 30 Han Chinese individuals (14 HSR patients and 16 controls) in our cohort. LTA toxicity greater than 12.5% ± 2.5% was considered positive. Differences among groups were determined by analysis of variance. In addition, we measured cytokine secretion in the patient sera between 1 month and 3 years after the event. All Han Chinese individuals and 30% of Caucasians were genotyped for HLA-B∗1502. A perfect correlation (r = 0.92) was observed between positive LTA and clinical diagnosis of DILI and SJS/toxic epidermal necrolysis (TEN). HLA-B∗1502 positivity in Han Chinese is a predictor of CBZ-HSR and PHY-HSR. HLA-B∗1502-negative Han Chinese receiving only CBZ or a combination of CBZ and PHY tolerated the drug(s) clinically, presenting negative CBZ-LTA and PHY-LTA. However, 3 patients presenting negative CBZ-LTA and PHY-LTA, as well as negative HLA-B∗1502, showed positive LTG-LTA (38%, 28%, and 25%, respectively), implying that they should not be prescribed LTG. Three patients had LTA positive to both PHY and CBZ, and 3 others had LTA positive to both PHY and LTG. Clinically, all 6 patients presented HSR to both drugs that they tested positive to (cross-reactivity). Patients were grouped based on the clinical presentation of their symptoms as only rash and fever or as a triad of rash, fever and DILI or SJS/TEN that characterizes “true” HSR. Levels of proinflammatory cytokines were significantly higher in patient sera compared with control sera. More specifically, the highest levels of tumor necrosis factor-α have been measured in patients presenting “true” HSR, as were the apoptotic markers Fas, caspase 8 activity, and M30. The LTA is sensitive for DILI and SJS/TEN regardless of drug or patient ethnicity. HSR prediction will prevent AED-induced morbidity. In Han Chinese, HLA-B∗1502 positivity is a predictor for CBZ-HSR and PHY-HSR. Its negativity does not predict a negative LTG-HSR. There is cross-reactivity between AEDs. Additionally, T-cell cytokines and chemokines control the pathogenesis of SJS/TEN and DILI, contributing to apoptotic processes in the liver and in the skin.

Dopamine receptor D1 mediates the inhibition of dopamine on the distal colonic motility

2012-01-27

The motility of distal colon could be inhibited by dopamine (DA), yet, the involved receptor is controversial according to the published reports. The goal of present study was to investigate DA receptor(s) mediated inhibition of DA on the colonic motility in rat. The contraction of isolated colon strips was assessed through isometric force transducer. The expressions of DA receptors in distal colon were detected through immunofluorescence and Western blot. DA concentration in colonic smooth muscle was measured by liquid chromatography/mass spectrometry. The results showed that DA inhibited the spontaneous motility of distal colon in a dose-dependent manner with EC50 8.3 μM. Tetrodotoxin increased colonic contractive frequency, but failed to affect the inhibition of DA on the colonic motility. Pretreatment with SCH-23390, an antagonist of dopaminergic receptor D1, shifted the dose-response curve to the right with EC50 of DA 37 μM. However, blocking dopaminergic receptor D2 with sulpiride, had no effect. The immunoreactivity of D1 and D2 were detected in the distal colon including myenteric plexus and smooth muscle. Acute cold-restraint stress (CRS) enhanced spontaneous contraction of rat distal colon, which was more sensitive to DA compared with control. Moreover, DA content and D1 expression in smooth muscle layer were increased under CRS condition. In conclusion, D1 in the smooth muscle is mediated DA inhibition on the spontaneous contraction of rat distal colon. The increased DA content and D1 receptor expression in the smooth muscle layer could be a compensatory effect under CRS condition to balance the enhanced colonic motility.

Intracardiac T-wave alternans, ischemia, and arrhythmias, in a canine model

John E. Madias — 2011-12-12

I enjoyed reading the article by Kwofie et al in the November 2011 issue of Translational Research, pertaining to the employment of intracardiac bipolar electrograms (IBEs)-based T-wave alternans (TWA) and ventricular tachycardia/fibrillation (VT/VF) in a canine ischemic model. The authors were careful to measure IBEs from both the risk zone and the surrounding normal sites, although I am puzzled about the use of drop in the QRS voltage in IBEs, as proof of myocardial ischemia (MI), considering that sometimes an increase in the amplitude of R-waves and T-waves is observed in the hyperacute phase of MI. However, there is backing from the literature regarding this concept, but still I am curious whether ST-segment elevation or depression or any changes in the T-wave polarity was detected with the emergence of MI. The authors found a correlation between TWA and MI, and an increase in the number of sites, magnitude, and discordance of TWA immediately preceding VT/VF. Also, as shown previously in normal human subjects, TWA in the canine model of this study was observed even before the generation of MI. The finding of the reversibility of TWA is intriguing and deserves further study. I will appreciate the response of the authors on the following: (1) in scrutinizing panels A and B of Fig 1, one can immediately appreciate that the QRS duration of EN1, EN2, EP3, and EN3 is wider in B than in A (after the imposition of MI); (2) this is particularly supported by the fact that the recording in B is slower because the horizontal time calibration bar is the same in length in A and B, and it represents 300 ms in A and 400 ms in B; (3) this implies that the change in TWA magnitude from A to B may be a result of the secondary changes of the T-waves resulting from the corresponding QRS complex changes; and (4) if this the case, the authors need to deal with the confounding factor of the increasing QRS duration in diagnosing and measuring TWA changes.

Erratum

2012-02-24

Dongsheng Fei, Xianglin Meng, Mingran Zhao, Kai Kang, Gang Tan, Shangha Pan, Yunpeng Luo, Wen Liu, Chuanchuan Nan, Hongchi Jiang, Geoffrey W Krissansen, Mingyan Zhao, Xueying Sun

Appreciation to Reviewers

Jeffrey Laurence, Michael Franklin — 2012-03-12

We wish to acknowledge the outstanding contribution of our reviewers and Editorial Advisory Board. The quality and breadth of the Journal is only made possible by the dedicated efforts of our reviewers.

Information for Readers

2012-05-01

Masthead

2012-05-01

Editorial Advisory Board

2012-05-01

Author Guidelines

2012-05-01

Translational Research

Contents

Role of advanced glycation endproducts and glyoxalase I in diabetic peripheral sensory neuropathy

Molecular profiling of individual tumor cells by hyperspectral microscopic imaging

Genetic analysis of the promoter region of the GATA4 gene in patients with ventricular septal defects

Uncoupling protein-2 expression and effects on mitochondrial membrane potential and oxidant stress in heart tissue

Chronic kidney disease as a risk factor for acute coronary syndromes in patients presenting to the emergency room with chest pain

Genetic and immune predictors for hypersensitivity syndrome to antiepileptic drugs

Dopamine receptor D1 mediates the inhibition of dopamine on the distal colonic motility

Intracardiac T-wave alternans, ischemia, and arrhythmias, in a canine model

Erratum

Appreciation to Reviewers

Information for Readers

Contents

Masthead

Editorial Advisory Board

Author Guidelines