Type 1 diabetes affects topoisomerase I activity and GlcNAcylation in rat organs: kidney, liver and pancreas.

Chronic hyperglycemia leads to the development of diabetes-induced organ complications, through changes in gene expression and protein function. We previously showed that in cell lines, topoisomerase I (topo I) is modified by O-GlcNAcylation, which affects its DNA relaxation activity. Since topo I participates in gene expression processes, we assumed that high glucose levels will affect its regulation and activity. Here we examined the effect of hyperglycemia on the regulation, GlcNAcylation and activity of topo I, in various internal rat organs that were subjected to diabetes-induced complications. Type 1 diabetes was induced in female rats by Streptozotocin injection. Topo I activity in nuclear protein extracts derived from diabetic and nondiabetic rat organs and topo I mRNA level were examined. Topo I and O-linked beta-N-acetylglucosamine (O-GlcNAc) transferase proteins and their O-GlcNAcylation were determined by western blot and immunoprecipitation assays. We show that topo I activity and enzyme protein level decreased in various tissues derived from the diabetic animals, whereas the enzyme mRNA level was not altered. Topo I protein was modified in vivo by O-GlcNAc, and O-GlcNAc transferase was coprecipitated with topo I protein, suggesting a possible interaction between both enzymes. This study demonstrates, for the first time, that topo I activity is regulated by high glucose levels, as a result of the diabetic state and is modified in vivo by O-GlcNAcylation, suggesting that topo I, an essential enzyme for gene expression, is involved in cellular processes which may lead to the pathogenesis of diabetic complications.

IMPA1 is essential for embryonic development and lithium-like pilocarpine sensitivity.

Lithium has been the standard pharmacological treatment for bipolar disorder over the last 50 years; however, the molecular targets through which lithium exerts its therapeutic effects are still not defined. We characterized the phenotype of mice with a dysfunctional IMPA1 gene (IMPA1-/-) to study the in vivo physiological functions of IMPA1, in general, and more specifically its potential role as a molecular target in mediating lithium-dependent physiological effects. Homozygote IMPA1-/- mice died in utero between days 9.5 and 10.5 post coitum (p.c.) demonstrating the importance of IMPA1 in early embryonic development. Intriguingly, the embryonic lethality could be reversed by myo-inositol supplementation via the pregnant mothers. In brains of adult IMPA1-/- mice, IMPase activity levels were found to be reduced (up to 65% in hippocampus); however, inositol levels were not found to be altered. Behavioral analysis of the IMPA1-/- mice indicated an increased motor activity in both the open-field test and the forced-swim test as well as a strongly increased sensitivity to pilocarpine-induced seizures, the latter supporting the idea that IMPA1 represents a physiologically relevant target for lithium. In conclusion the IMPA1-/- mouse represents a novel model to study inositol homeostasis, and indicates that genetic inactivation of IMPA1 can mimic some actions of lithium.

Response of Symptomatic Persistent Chronic Disseminated Candidiasis to Corticosteroid Therapy in Immunosuppressed Pediatric Patients: Case Study and Review of the Literature.

BACKGROUND: Chronic disseminated candidiasis (CDC) is a severe invasive fungal infection principally observed during neutrophil recovery in patients with acute leukemia treated with intensive chemotherapy. Its pathophysiology remains unclear. We describe the management of 6 children with symptomatic CDC who did not respond to antifungal therapy. METHODS: The databases of the hematology-oncology departments of 2 tertiary pediatric medical centers were searched for all patients diagnosed with CDC from 2003 to 2015, who responded to corticosteroids after failing antifungal therapy. Clinical, laboratory and radiologic data were collected. RESULTS: Six patients (3 women, 3 men; 9-18 years of age) met the study criteria. Underlying diseases were acute lymphoblastic leukemia (n = 3) and large B-cell lymphoma, acute myeloid leukemia and severe aplastic anemia (n = 1 each). Presenting symptoms/signs of CDC were fever in all cases, with abdominal or chest pain, and/or mucositis. Candida infection was identified in blood cultures in 4 patients and in bronchoalveolar lavage fluid in one. In the absence of response to antifungal agents (4-50 days from CDC diagnosis), prednisone 2 mg/kg/day or equivalent was administered. CDC-attributable clinical symptoms resolved in 4 patients within 6-19 days; one patient required an additional nonsteroidal anti-inflammatory agent. Abnormalities on imaging decreased or disappeared within 5 months to 2 years in 4 patients. CONCLUSIONS: In children with persistent symptomatic CDC, despite adequate antifungal therapy, administration of corticosteroids may yield rapid resolution of symptoms and decreased inflammatory markers. In patients who do not respond to steroids, the addition of a nonsteroidal anti-inflammatory drug should be considered.

Postmortem parietal cortex TPH2 expression is not altered in schizophrenic, unipolar-depressed, and bipolar patients vs control subjects.

Serotonin (5-hydroxytryptamine [5-HT]) is a neurotransmitter synthesized in the raphe nuclei of the brain stem in the central nervous system (CNS) and also in the periphery. Dysfunction of the serotonergic system has been implicated in the pathogenesis of psychiatric disorders. Tryptophan hydroxylase (TPH) is the rate-limiting enzyme in 5-HT biosynthesis. For more than a decade, only one gene encoding TPH was identified in vertebrates. Recently, a second TPH gene, designated TPH2, was detected, located on human chromosome 12, a susceptibility region for affective disorders. TPH2 is predominantly expressed in the brain, whereas the classical TPH gene, TPH1, is expressed in peripheral tissues. The discovery of the brain-abundant TPH2 gene justifies a new concept of the CNS serotonergic system. TPH2, rather than TPH1, has now become a candidate gene for 5-HT-related affective disorders. We compared TPH2 mRNA levels in postmortem parietal cortex of unipolar-depressed, bipolar, and schizophrenic patients vs control subjects, using real-time reverse transcription polymerase chain reaction. No significant difference in TPH2 mRNA levels was found among the four diagnostic groups. The lack of difference might suggest that this gene is not involved in the etiology of of these psychiatric disorders. Alternatively, it is possible that the parietal cortex is not the relevant brain area involved in the pathophysiology of these disorders or that posttranscriptional modifications of TPH2 mRNA occur in these patients, causing changes in protein levels and/or enzymatic activity.

Health provider determinants of nonattendance in pediatric otolaryngology patients.

INTRODUCTION: Nonattendance for otolaryngology appointments disrupts the management of medical care and leads to ineffective use of resources. The determinants of nonattendance in pediatric otolaryngology patients have not been well documented. OBJECTIVES: To investigate health provider determinants of nonattendance in pediatric otolaryngology patients. STUDY DESIGN: We assessed the effects of waiting time for an appointment and the timing of the appointment (during the day, week, and year) on nonattendance proportions during a 1 year period. Chi square tests were used to analyze statistically significant differences of categorical variables. Logistic regression was used for multivariate analyses. RESULTS: A total of 2,628 pediatric visits were included in the study. The overall proportion of nonattendance at the pediatric otolaryngology clinic was 33.0%. Nonattendance proportions were 32.7% between 7 AM and 9 AM; 28.3% between 9 AM and 2 PM, and 36.5% between 2 PM and 8 PM (P < .001). The proportion of nonattendance was 24.1% when there was a short waiting time for an appointment (0-7 days), and 36.3% when there was an intermediate waiting time (7-15 days), and 36.6% when there was a long waiting time (15 days and above)(P < .001, P < .012, respectively). CONCLUSIVE: Health provider determinants of nonattendance in pediatric otolaryngology clinic appointments include the waiting time for an appointment and the hour of the appointment within the day.

Long-term dietary changes after vertical banded gastroplasty: is the trade-off favorable?

BACKGROUND: Insufficient data exist about the long-term health consequences of gastric restriction procedures used for treatment of obesity. The long-term nutritional changes that occur after vertical banded gastroplasty (VBG) were evaluated. METHODS: All consecutive patients who underwent VBG surgery in one surgical ward were invited for a follow-up study 3-10 years after the surgery. Demographic and clinical characteristics were evaluated from the patients' medical charts. Dietary assessment was performed using a food frequency questionnaire, which included 52 frequently consumed food items. RESULTS: Of the 122 patients who underwent VBG between 1986-1992, 75 (62%) participated in the follow-up study. The average time since surgery was 5.4 +/- 1.8 years, and the average weight loss was 24.9 +/- 12.4%. Most of the patients eat only one major meal daily, and only one-third regularly ingest solid foods. Dietary analysis revealed a decreased intake of most nutrients compared with pre-surgery, with the exception of dairy products, sweet foods and fluids. The greatest decrease was found in the consumption of fiber-rich fruits and vegetables, followed by meat, fish and complex carbohydrates. CONCLUSION: While the weight loss itself and the reduction in fat consumption that are seen after VBG are probably beneficial, the long-term effects of the decreased consumption of fruit, vegetables, other complex carbohydrates and fish may counterbalance these benefits. The net effect of this trade-off on future health is difficult to predict and requires long-term evaluation of clinical outcome.

Determinants of long-term satisfaction after vertical banded gastroplasty.

BACKGROUND: The long-term usefulness of vertical banded gastroplasty (VBG) in achieving weight loss is controversial, and adverse effects related to the procedure may attenuate patient satisfaction. Our objective was to evaluate patient satisfaction, and to identify parameters that are related to such satisfaction, 3 to10 years after VBG. METHODS: All consecutive patients who underwent VBG in one surgical ward were invited for a follow-up study 3 to 10 years after surgery. Questions relating to symptoms and quality of life were evaluated in a personal interview using a structured questionnaire. RESULTS: Of the 122 patients who underwent VBG from 1986 to 1992, 75 patients were located and agreed to participate in the follow-up study. The average time since surgery was 5.4 +/- 1.8 years. The average weight loss was 24.9 +/- 12.4%, representing an excess body-weight loss of 58.6 +/- 30.4%. Overall, 65% of the patients were satisfied with the results of surgery while 19% expressed dissatisfaction. Significant improvement was seen in respiratory difficulties, ability to perform physical exercise, and mental status. Successful weight loss and the frequency of respiratory difficulties were the only independent parameters associated with patient satisfaction. Although vomiting, gastroesophageal reflux and difficulty in swallowing occurred in over two-thirds of the patients, their presence was not correlated with patient dissatisfaction. CONCLUSION: Despite the presence of a multitude of adverse effects, the majority of our patients were satisfied with the long-term results of VBG. Successful weight loss and improvement in respiratory difficulties were the major determinants of patient satisfaction.

Inhibition of monocyte chemotaxis by VB-201, a small molecule lecinoxoid, hinders atherosclerosis development in ApoE⁻/⁻ mice.

OBJECTIVE: Monocytes are motile cells which sense inflammatory stimuli and subsequently migrate to sites of inflammation. Key players in host defense, monocytes have nevertheless been implicated as requisite mediators of several chronic inflammatory diseases. Inhibition of monocyte chemotaxis is therefore an attractive anti-inflammatory strategy. Oxidized phospholipids (OxPL) are native regulators of inflammation, yet their direct effect on monocyte chemotaxis is poorly defined. In this study, we investigated the direct effect of natural and synthetic phospholipids on monocyte chemotaxis. METHODS: Exploring various phospholipids using in vitro chemotaxis assays, we found that the natural phospholipid 1-palmitoyl-2-glutaryl phosphatidylcholine (PGPC) can decrease monocyte chemotaxis by 50%, while other tested OxPL had no effect. We generated a library of synthetic OxPL designated lecinoxoids, which was screened for anti-inflammatory properties. RESULTS AND CONCLUSIONS: VB-201, a small-molecule lecinoxoid, exhibited up to 90% inhibition of monocyte chemotaxis in vitro. Molecular analysis revealed that the effect of VB-201 was not restricted to a specific chemotactic ligand or receptor, and resulted from inhibition of signaling pathways required for monocyte chemotaxis. Interestingly, VB-201 did not inhibit monocyte adhesion or phagocytosis and had no effect on chemotaxis of CD4(+) T-cells or neutrophils. In vivo, oral treatment with VB-201 reduced monocyte migration in a peritonitis model and inhibited atheroma development in ApoE(-/-) mice, without affecting cholesterol or triglyceride levels. Our findings highlight a novel role played by native and synthetic phospholipids in regulation of monocyte chemotaxis. The data strengthen the involvement of phospholipids as key signaling molecules in inflammatory settings and demonstrate their potential therapeutic applicability.

Inhibition of inositol monophosphatase (IMPase) at the calbindin-D28k binding site: molecular and behavioral aspects.

Bipolar-disorder (manic-depressive illness) is a severe chronic illness affecting ∼1% of the adult population. It is treated with mood-stabilizers, the prototypic one being lithium-salts (lithium), but it has life threatening side-effects and a significant number of patients fail to respond. The lithium-inhibitable enzyme inositol-monophosphatase (IMPase) is one of the viable targets for lithium's mechanism of action. Calbindin-D28k (calbindin) up-regulates IMPase activity. The IMPase-calbindincomplex was modeled using the program MolFit. The in-silico model indicated that the 55-66 amino-acid segment of IMPase anchors calbindin via Lys59 and Lys61 with a glutamate in between (Lys-Glu-Lys motif) and that the motif interacts with residues Asp24 and Asp26 of calbindin. We found that differently from wildtype calbindin, IMPase was not activated by mutated calbindin in which Asp24 and Asp26 were replaced by alanine. Calbindin's effect was significantly reduced by a linear peptide with the sequence of amino acids 58-63 of IMPase (peptide 1) and by six amino-acid linear peptides including at least part of the Lys-Glu-Lys motif. The three amino-acid peptide Lys-Glu-Lys or five amino-acid linear peptides containing this motif were ineffective. Mice administered peptide 1 intracerebroventricularly exhibited a significant anti-depressant-like reduced immobility in the forced-swim test. Based on the sequence of peptide 1, and to potentially increase the peptide's stability, cyclic and linear pre-cyclic analog peptides were synthesized. One cyclic peptide and one linear pre-cyclic analog peptide inhibited calbindin-activated brain IMPase activity in-vitro. Our findings may lead to the development of molecules capable of inhibiting IMPase activity at an alternative site than that of lithium.

Modification of topoisomerase I activity by glucose and by O-GlcNAcylation of the enzyme protein.

The regulation of topoisomerase I (topo I) activity is of prime importance for gene expression. It participates in DNA replication, transcription, recombination, and DNA repair, and serves as a target for anticancer drugs. Many proteins and enzymes are modified by O-linked beta-N-acetylglucosamine (O-GlcNAc), which exerts profound effects on their function. However, the modification of topo I by O-GlcNAc and the effect on its activity has not been previously reported. Here, we show that topo I protein is modified by O-GlcNAc in vitro in the porcine proximal tubular epithelial cell line (LLPCK-1), and in vivo in the mouse kidney. The level of O-GlcNAcylation of topo I protein correlates well with the enzyme activity, namely, a decrease in O-GlcNAc results in a reduction in topo I activity, and vice versa. O-GlcNAc transferase (OGT) was coprecipitated with topo I protein, suggesting a possible interaction between both enzymes. In addition, treatment of cells with glucosamine increased topo I activity and O-GlcNAcylation. The results of this study provide a novel mechanism for the regulation of topo I activity. Topo I is important for DNA transcription, therefore, its regulation by GlcNAcylation contributes to the mechanism by which glucose levels affect gene expression, and may pave the way to the development of new drugs that could control topo I activity.

Health provider factors associated with nonattendance in pediatric dermatology ambulatory patients.

Nonattendance for dermatology appointments disrupts the management of medical delivery and leads to inefficient allocation of resources and lost revenue. The factors that determine nonattendance in pediatric dermatology patients have not been well documented. We investigated health provider factors for nonattendance in pediatric dermatology patients. We assessed the effects of waiting time for an appointment and the timing of the appointment (during the day, week, and year) on nonattendance proportions during a 1 year period. Chi-squared tests were used to analyze statistically significant differences of categorical variables. Logistic regression was used for multivariate analyses. A total of 1696 children visits were included in the study. The overall rate of nonattendance at the dermatology clinic was 30.5%. Nonattendance was 29.7% during the periods between 8 a.m. to 1 p.m. and 3 p.m. to 7 p.m. and 40.7% during the periods between 1 p.m. to 3 p.m. and 7 p.m. to 8 p.m. (p = 0.013). Nonattendance was 21.1% when the waiting time for an appointment was short (1-7 days), 32.5% when it was intermediate (8-14 days) and 43.5% when the wait time was long (15 days and above) (p-value < 0.001). A multivariate logistic regression model demonstrated that the hour of the day and the waiting time for an appointment were significantly associated with nonattendance (p value = 0.009, p value < 0.001, respectively). We conclude that in children attending a dermatology clinic, health provider factors that determine nonattendance include the waiting time for an appointment and the hour of the appointment within the day.

Lymphocyte G-protein receptor kinase (GRK)3 mRNA levels in bipolar disorder.

Linkage studies in bipolar disorder were positive for markers in the region of chromosome 22q12.1 including the gene coding for G-protein receptor kinase (GRK)3. Two of six variants of the GRK3 5'-UTR/promoter were reported to be associated with bipolar disorder. GRK3 protein levels in lymphoblastoid cell lines derived from bipolar patients originating from families with linkage to chromosome 22q11 were reported to be decreased compared to those of control subjects and correlated with disease severity. We compared GRK3 mRNA levels in fresh lymphocytes from 31 bipolar patients vs. 26 control subjects, using real-time RT-PCR. No overall difference was found between patients and controls. However, GRK3 mRNA levels were markedly and significantly reduced in the subgroup of patients with no family history of a major psychiatric disorder compared with patients with family history.

Biological Warfare: Implications for Antimicrobial Use.

Biological warfare is intended to incapacitate a large number of individuals at a single exposure, creating epidemic-type disease, death, and social chaos. The organisms with potential for immediate use as bacteriologic weapons are Bacillus anthracis, Brucella melitensis, Yersinia pestis, and Vibrio cholera, all necessitating antibiotic therapy for a cure. It is reasonable, therefore, to assume that a biological attack, or even a hoax, would requiure thousands of individuals over a large area to begin antibiotic therapy. Issues such as antibiotic availability, logistical problems in antibiotic distribution, development of drug resistance, side effects influencing the individual, and adverse effects on the community due to the impact of mass therapy on the ecology, make biological warfare the most apocalyptic scenario for the creation of a "postantibiotic era."

Severe Coxsackie virus B infection in preterm newborns treated with pleconaril.

UNLABELLED: Four preterm newborn infants with severe multisystem Coxsackie virus B infection were treated with an oral suspension of pleconaril (5 mg/kg per day). The patients had myocarditis, fulminant hepatitis, meningoencephalitis and disseminated intravascular coagulopathy. All four infants recovered, and no adverse effects of the treatment were noted. CONCLUSION: pleconaril needs to be comprehensively evaluated in this population.