Subchronic oral toxicity study of decitabine in combination with tetrahydrouridine in CD-1 mice.

Decitabine (5-aza-2'-deoxycytidine; DAC) in combination with tetrahydrouridine (THU) is a potential oral therapy for sickle cell disease and ß-thalassemia. A study was conducted in mice to assess safety of this combination therapy using oral gavage of DAC and THU administered 1 hour prior to DAC on 2 consecutive days/week for up to 9 weeks followed by a 28-day recovery to support its clinical trials up to 9-week duration. Tetrahydrouridine, a competitive inhibitor of cytidine deaminase, was used in the combination to improve oral bioavailability of DAC. Doses were 167 mg/kg THU followed by 0, 0.2, 0.4, or 1.0 mg/kg DAC; THU vehicle followed by 1.0 mg/kg DAC; or vehicle alone. End points evaluated were clinical observations, body weights, food consumption, clinical pathology, gross/histopathology, bone marrow micronuclei, and toxicokinetics. There were no treatment-related effects noticed on body weight, food consumption, serum chemistry, or urinalysis parameters. Dose- and gender-dependent changes in plasma DAC levels were observed with a Cmax within 1 hour. At the 1 mg/kg dose tested, THU increased DAC plasma concentration (∼ 10-fold) as compared to DAC alone. Severe toxicity occurred in females receiving high-dose 1 mg/kg DAC + THU, requiring treatment discontinuation at week 5. Severity and incidence of microscopic findings increased in a dose-dependent fashion; findings included bone marrow hypocellularity (with corresponding hematologic changes and decreases in white blood cells, red blood cells, hemoglobin, hematocrit, reticulocytes, neutrophils, and lymphocytes), thymic/lymphoid depletion, intestinal epithelial apoptosis, and testicular degeneration. Bone marrow micronucleus analysis confirmed bone marrow cytotoxicity, suppression of erythropoiesis, and genotoxicity. Following the recovery period, a complete or trend toward resolution of these effects was observed. In conclusion, the combination therapy resulted in an increased sensitivity to DAC toxicity correlating with DAC plasma levels, and females are more sensitive compared to their male counterparts.

Can harmonic focus curved shear effectively seal the pancreatic ducts and prevent pancreatic leak? Feasibility evaluation and testing in ex vivo and in vivo porcine models.

BACKGROUND: The purpose of this study was to evaluate the ability of Harmonic energy technology to transect and seal the pancreatic duct compared with the standard monopolar electrosurgery transection and oversew technique in a porcine distal pancreatectomy survival model. Harmonic energy technology is as effective as standard oversew technique for preventing pancreatic leak after distal pancreatectomy. METHODS: The animal protocol used for this study was approved by the Institutional Animal Care and Use Committee (IACUC) prior to the conduct of the study. Spleen-preserving distal pancreatectomy was performed in seven pigs (80-100 lb) by the same surgeon. In four animals, the pancreas was divided with the Harmonic Focus Curved Shears (test group) with no additional suturing for control of hemostasis or leak. In three animals, the pancreas was divided using monopolar electrosurgery (30 W coagulation) and the cut end of the pancreas was oversewn with a locking suture (control group). A previously worked out standard operative technique was used in all procedures. Operating end points included surgery time and blood loss. Animals were euthanized and necropsied at 7 to 8 d following surgery. Survival endpoints included clinical response to surgery, serum chemistry profiles before surgery and at necropsy, and histology of the pancreas transection site. RESULTS: Mean operative time for pancreatic resection was 15min in the control group and 10min in the test group. No significant blood loss was noted in either group. The median size of the resected pancreas was 4.3 cm. Three animals in the control group and three in the test group completed the study without complications. One animal in the test group failed to eat, appeared dehydrated, and was taken off study on POD#2. In this animal, there was a doubling of serum lipase at euthanasia and gross evidence of ileus, which was attributed to a pancreatic leak. Histologic examination of the residual pancreas in both groups at necropsy revealed a 2-5 mm band of necrotic tissue associated with neutrophilic infiltration in the control group, and less than a 1mm band of necrotic tissue in the test group. CONCLUSION: The Harmonic Focus Curved Shears, using Harmonic energy technology, appears to seal the pancreatic ducts and prevent pancreatic leak at 75% (3/4) efficiency in this survival model. However, the survival leak rate, 25% (1/4) was higher in the Harmonic Focus Curved Shears test group compared with the control oversewn group 0% (0/3). This feasibility study shows potential for Harmonic technology to be used to seal the pancreatic ducts, but additional testing and optimization of surgical techniques are needed.

Chronic inhalation of short asbestos: lung fiber burdens and histopathology for monkeys maintained for 11.5 years after exposure.

In an earlier report, Platek et al. (1985) presented the results of an 18-month inhalation exposure of rats and monkeys to short chrysotile asbestos. The mean chamber exposure level was 1.0 mg/m(3) with an average of 0.79 fibers/ml > 5 microm in length. Gross and histopathological examination of exposed and control rats indicated no treatment-related lesions. Asbestos bodies adjacent to the terminal bronchioles, but no fibrosis, were found in lung biopsy tissue taken from the exposed monkeys at 10 months post-exposure. Fifteen monkeys (9 exposed and 6 controls) from this study were maintained for 11.5 years following exposure. Lung fiber burdens were determined by transmission electron microscopy. The mean lung burden (+/- standard deviation) for 59 samples from exposed monkeys was 63 +/- 30 x 10(6) fibers/g dry lung (range, 18-139 x 10(6)). The geometric mean fiber length was 3.5 microm with 35% of the fibers being > 5 microm in length. These data indicate some chrysotile fibers are durable in vivo for a significant period of time. Lungs were examined grossly and microscopically. No lesions attributable to the inhalation exposure were noted. Asbestos bodies were seen in the lungs of treated monkeys, primarily in the interstitium near bronchioles or small pulmonary blood vessels (which also may have been near to bronchioles just out of the plane of section).

Global Transcriptome Analysis of RNA Abundance Regulation by ADAR in Lung Adenocarcinoma.

Despite tremendous advances in targeted therapies against lung adenocarcinoma, the majority of patients do not benefit from personalized treatments. A deeper understanding of potential therapeutic targets is crucial to increase the survival of patients. One promising target, ADAR, is amplified in 13% of lung adenocarcinomas and in-vitro studies have demonstrated the potential of its therapeutic inhibition to inhibit tumor growth. ADAR edits millions of adenosines to inosines within the transcriptome, and while previous studies of ADAR in cancer have solely focused on protein-coding edits, >99% of edits occur in non-protein coding regions. Here, we develop a pipeline to discover the regulatory potential of RNA editing sites across the entire transcriptome and apply it to lung adenocarcinoma tumors from The Cancer Genome Atlas. This method predicts that 1413 genes contain regulatory edits, predominantly in non-coding regions. Genes with the largest numbers of regulatory edits are enriched in both apoptotic and innate immune pathways, providing a link between these known functions of ADAR and its role in cancer. We further show that despite a positive association between ADAR RNA expression and apoptotic and immune pathways, ADAR copy number is negatively associated with apoptosis and several immune cell types' signatures.

Triclosan Induces Thymic Stromal Lymphopoietin in Skin Promoting Th2 Allergic Responses.

Triclosan is an antimicrobial chemical incorporated into many personal, medical and household products. Approximately, 75% of the U.S. population has detectable levels of triclosan in their urine, and although it is not typically considered a contact sensitizer, recent studies have begun to link triclosan exposure with augmented allergic disease. We examined the effects of dermal triclosan exposure on the skin and lymph nodes of mice and in a human skin model to identify mechanisms for augmenting allergic responses. Triclosan (0%-3%) was applied topically at 24-h intervals to the ear pinnae of OVA-sensitized BALB/c mice. Skin and draining lymph nodes were evaluated for cellular responses and cytokine expression over time. The effects of triclosan (0%-0.75%) on cytokine expression in a human skin tissue model were also examined. Exposure to triclosan increased the expression of TSLP, IL-1ß, and TNF-α in the skin with concomitant decreases in IL-25, IL-33, and IL-1α. Similar changes in TSLP, IL1B, and IL33 expression occurred in human skin. Topical application of triclosan also increased draining lymph node cellularity consisting of activated CD86(+)GL-7(+) B cells, CD80(+)CD86(+) dendritic cells, GATA-3(+)OX-40(+)IL-4(+)IL-13(+) Th2 cells and IL-17 A(+) CD4 T cells. In vivo antibody blockade of TSLP reduced skin irritation, IL-1ß expression, lymph node cellularity, and Th2 responses augmented by triclosan. Repeated dermal exposure to triclosan induces TSLP expression in skin tissue as a potential mechanism for augmenting allergic responses.

Leucine-rich repeat kinase 2 (LRRK2)-deficient rats exhibit renal tubule injury and perturbations in metabolic and immunological homeostasis.

Genetic evidence links mutations in the LRRK2 gene with an increased risk of Parkinson's disease, for which no neuroprotective or neurorestorative therapies currently exist. While the role of LRRK2 in normal cellular function has yet to be fully described, evidence suggests involvement with immune and kidney functions. A comparative study of LRRK2-deficient and wild type rats investigated the influence that this gene has on the phenotype of these rats. Significant weight gain in the LRRK2 null rats was observed and was accompanied by significant increases in insulin and insulin-like growth factors. Additionally, LRRK2-deficient rats displayed kidney morphological and histopathological alterations in the renal tubule epithelial cells of all animals assessed. These perturbations in renal morphology were accompanied by significant decreases of lipocalin-2, in both the urine and plasma of knockout animals. Significant alterations in the cellular composition of the spleen between LRRK2 knockout and wild type animals were identified by immunophenotyping and were associated with subtle differences in response to dual infection with rat-adapted influenza virus (RAIV) and Streptococcus pneumoniae. Ontological pathway analysis of LRRK2 across metabolic and kidney processes and pathological categories suggested that the thioredoxin network may play a role in perturbing these organ systems. The phenotype of the LRRK2 null rat is suggestive of a complex biology influencing metabolism, immune function and kidney homeostasis. These data need to be extended to better understand the role of the kinase domain or other biological functions of the gene to better inform the development of pharmacological inhibitors.