Vasopressin-induced hyponatremia in an adult normotensive trauma patient: A case report.

BACKGROUND: Arginine vasopressin is a neuropeptide produced in the hypothalamus and released by the posterior pituitary gland. In addition to maintaining plasma osmolarity, under hypovolemic or hypotensive conditions, it helps maintain plasma volume through renal water reabsorption and increases systemic vascular tone. Its synthetic analogues are widely used in the intensive care unit as a continuous infusion, in addition to hospital floors as an intravenous or intranasal dose. A limited number of cases of hyponatremia in patients with septic or hemorrhagic shock have been reported previously with vasopressin. We report for the first time a normotensive patient who developed vasopressin-induced hyponatremia. CASE SUMMARY: A 39-year-old man fell off a forklift and sustained an axial load injury to his cranium. He had no history of previous trauma. Examination was normal except for motor and sensory deficits. The Imagine test showed endplate fracture at C7 and acute traumatic disc at C7 with cortical degeneration. He underwent cervical discectomy and fusion, laminectomy, and posterior instrumented fusion. After intensive care unit admission post-surgery, he developed hyponatremia of 121-124 mEq/L post phenylephrine and vasopressin infusion to maintain blood pressure maintenance. He was evaluated for syndrome of inappropriate secretion of antidiuretic hormone, hypothyroid, adrenal-induced, or diuretic-induced hyponatremia. At the end of extensive evaluation for the underlying cause of hyponatremia, vasopressin was discontinued. He was also put on fluid restriction, given exogenous desmopressin, and a dextrose 5% in water infusion to prevent osmotic demyelination syndrome caused by sodium overcorrection which improved his sodium level to 135 mmol/L. CONCLUSION: The presentation of vasopressin-induced hyponatremia is uncommon in normotensive patients, and the most difficult aspect of this condition is determining the underlying cause of hyponatremia. Our case illustrates that, considering the vast differential diagnosis of hyponatremia in hospitalized patients, both hospitalists and intensivists should be aware of this serious complication of vasopressin therapy.

Combined targeting of soluble latent TGF-ß and a solid tumor-associated antigen with adapter CAR T cells.

Solid tumors consist of malignant and nonmalignant cells that together create the local tumor microenvironment (TME). Additionally, the TME is characterized by the expression of numerous soluble factors such as TGF-ß. TGF-ß plays an important role in the TME by suppressing T cell effector function and promoting tumor invasiveness. Up to now CAR T cells exclusively target tumor-associated antigens (TAA) located on the cell membrane. Thus, strategies to exploit soluble antigens as CAR targets within the TME are needed. This study demonstrates a novel approach using Adapter CAR (AdCAR) T cells for the detection of soluble latent TGF-ß within the TME of a pancreatic tumor model. We show that AdCARs in combination with the respective adapter can be used to sense soluble tumor-derived latent TGF-ß, both in vitro and in vivo. Sensing of the soluble antigen induced cellular activation and effector cytokine production in AdCAR T cells. Moreover, we evaluated AdCAR T cells for the combined targeting of soluble latent TGF-ß and tumor cell killing by targeting CD66c as TAA in vivo. In sum, our study broadens the spectrum of targetable moieties for AdCAR T cells by soluble latent TGF-ß.

VRK1 as a synthetic lethal target in VRK2 promoter-methylated cancers of the nervous system.

Collateral lethality occurs when loss of a gene/protein renders cancer cells dependent on its remaining paralog. Combining genome-scale CRISPR/Cas9 loss-of-function screens with RNA sequencing in over 900 cancer cell lines, we found that cancers of nervous system lineage, including adult and pediatric gliomas and neuroblastomas, required the nuclear kinase vaccinia-related kinase 1 (VRK1) for their survival in vivo. VRK1 dependency was inversely correlated with expression of its paralog VRK2. VRK2 knockout sensitized cells to VRK1 loss, and conversely, VRK2 overexpression increased cell fitness in the setting of VRK1 loss. DNA methylation of the VRK2 promoter was associated with low VRK2 expression in human neuroblastomas and adult and pediatric gliomas. Mechanistically, depletion of VRK1 reduced barrier-to-autointegration factor phosphorylation during mitosis, resulting in DNA damage and apoptosis. Together, these studies identify VRK1 as a synthetic lethal target in VRK2 promoter-methylated adult and pediatric gliomas and neuroblastomas.

Transition to a mesenchymal state in neuroblastoma confers resistance to anti-GD2 antibody via reduced expression of ST8SIA1.

Immunotherapy with anti-GD2 antibodies has advanced the treatment of children with high-risk neuroblastoma, but nearly half of patients relapse, and little is known about mechanisms of resistance to anti-GD2 therapy. Here, we show that reduced GD2 expression was significantly correlated with the mesenchymal cell state in neuroblastoma and that a forced adrenergic-to-mesenchymal transition (AMT) conferred downregulation of GD2 and resistance to anti-GD2 antibody. Mechanistically, low-GD2-expressing cell lines demonstrated significantly reduced expression of the ganglioside synthesis enzyme ST8SIA1 (GD3 synthase), resulting in a bottlenecking of GD2 synthesis. Pharmacologic inhibition of EZH2 resulted in epigenetic rewiring of mesenchymal neuroblastoma cells and re-expression of ST8SIA1, restoring surface expression of GD2 and sensitivity to anti-GD2 antibody. These data identify developmental lineage as a key determinant of sensitivity to anti-GD2 based immunotherapies and credential EZH2 inhibitors for clinical testing in combination with anti-GD2 antibody to enhance outcomes for children with neuroblastoma.

Genomic inverse PCR for exploration of ligated breakpoints (GIPFEL), a new method to detect translocations in leukemia.

Here we present a novel method "Genomic inverse PCR for exploration of ligated breakpoints" (GIPFEL) that allows the sensitive detection of recurrent chromosomal translocations. This technique utilizes limited amounts of DNA as starting material and relies on PCR based quantification of unique DNA sequences that are created by circular ligation of restricted genomic DNA from translocation bearing cells. Because the complete potential breakpoint region is interrogated, a prior knowledge of the individual, specific interchromosomal fusion site is not required. We validated GIPFEL for the five most common gene fusions associated with childhood leukemia (MLL-AF4, MLL-AF9, MLL-ENL, ETV6-RUNX1, and TCF3-PBX1). A workflow of restriction digest, purification, ligation, removal of linear fragments and precipitation enriching for circular DNA was developed. GIPFEL allowed detection of translocation specific signature sequences down to a 10-4 dilution which is close to the theoretical limit. In a blinded proof-of-principle study utilizing DNA from cell lines and 144 children with B-precursor-ALL associated translocations this method was 100% specific with no false positive results. Sensitivity was 83%, 65%, and 24% for t(4;11), t(9;11) and t(11;19) respectively. Translocation t(12;21) was correctly detected in 64% and t(1;19) in 39% of the cases. In contrast to other methods, the characteristics of GIPFEL make it particularly attractive for prospective studies.

Eyelid desmoplastic trichilemmoma: 2 case reports and review.

Desmoplastic trichilemmomas (DTs) are a rare variant of trichilemmomas first described in 1985. Since then, 11 cases involving the eyelid have been reported. Two cases of this study raise this total to 13. DTs are often clinically confused with basal cell carcinoma and must be proven by biopsy to obtain the correct diagnosis. These lesions are more commonly present in patients older than 50 years. Despite the fact that they are benign, they are often associated with basal cell carcinomas and as such should be managed carefully, requiring excisional biopsy with frozen borders or Mohs controlled margins.

Percutaneous embolization of an orbital hemangiopericytoma with Onyx facilitates its surgical excision.

This is a case report on the surgical treatment of orbital hemangiopericytoma. The patient initially underwent an attempted excision of the tumor with significant hemorrhage elsewhere, which precluded its complete excision. Due to continued growth and pain, the patient underwent percutaneous embolization with Onyx-18 under fluoroscopic radiographic guidance. The patient continued to have pain, progressive enlargement of the tumor, and underwent surgical excision. The excision was noted to have minimal bleeding and was relatively easy to dissect and excise compared with the typical hemangiopericytoma. Embolization with Onyx-18 may be a useful preoperative treatment option for hemangiopericytomas and other vascular lesions of the orbit, and elsewhere.

δ-Tocotrienol and quercetin reduce serum levels of nitric oxide and lipid parameters in female chickens.

BACKGROUND: Chronic, low-grade inflammation provides a link between normal ageing and the pathogenesis of age-related diseases. A series of in vitro tests confirmed the strong anti-inflammatory activities of known inhibitors of NF-κB activation (δ-tocotrienol, quercetin, riboflavin, (-) Corey lactone, amiloride, and dexamethasone). δ-Tocotrienol also suppresses ß-hydroxy-ß-methylglutaryl coenzyme A (HMG-CoA) reductase activity (the rate-limiting step in de novo cholesterol synthesis), and concomitantly lowers serum total and LDL cholesterol levels. We evaluated these compounds in an avian model anticipating that a dietary additive combining δ-tocotrienol with quercetin, riboflavin, (-) Corey lactone, amiloride, or dexamethasone would yield greater reductions in serum levels of total cholesterol, LDL-cholesterol and inflammatory markers (tumor necrosis factor-α [TNF-α], and nitric oxide [NO]), than that attained with the individual compounds. RESULTS: The present results showed that supplementation of control diets with all compounds tested except riboflavin, (-) Corey lactone, and dexamethasone produced small but significant reductions in body weight gains as compared to control. (-) Corey lactone and riboflavin did not significantly impact body weight gains. Dexamethasone significantly and markedly reduced weight gain (>75%) compared to control. The serum levels of TNF-α and NO were decreased 61% - 84% (P < 0.001), and 14% - 67%, respectively, in chickens fed diets supplemented with δ-tocotrienol, quercetin, riboflavin, (-) Corey lactone, amiloride, or dexamethasone as compared to controls. Significant decreases in the levels of serum total and LDL-cholesterol were attained with δ-tocotrienol, quercetin, riboflavin and (-) Corey lactone (13% - 57%; P < 0.05), whereas, these levels were 2-fold higher in dexamethasone treated chickens as compared to controls. Parallel responses on hepatic lipid infiltration were confirmed by histological analyses. Treatments combining δ-tocotrienol with the other compounds yielded values that were lower than individual values attained with either δ-tocotrienol or the second compound. Exceptions were the significantly lower total and LDL cholesterol and triglyceride values attained with the δ-tocotrienol/(-) Corey lactone treatment and the significantly lower triglyceride value attained with the δ-tocotrienol/riboflavin treatment. δ-Tocotrienol attenuated the lipid-elevating impact of dexamethasone and potentiated the triglyceride lowering impact of riboflavin. Microarray analyses of liver samples identified 62 genes whose expressions were either up-regulated or down-regulated by all compounds suggesting common impact on serum TNF-α and NO levels. The microarray analyses further identified 41 genes whose expression was differentially impacted by the compounds shown to lower serum lipid levels and dexamethasone, associated with markedly elevated serum lipids. CONCLUSIONS: This is the first report describing the anti-inflammatory effects of δ-tocotrienol, quercetin, riboflavin, (-) Corey lactone, amiloride, and dexamethasone on serum TNF-δ and NO levels. Serum TNF-δ levels were decreased by >60% by each of the experimental compounds. Additionally, all the treatments except with dexamethasone, resulted in lower serum total cholesterol, LDL-cholesterol and triglyceride levels. The impact of above mentioned compounds on the factors evaluated herein was increased when combined with δ-tocotrienol.

NCAM is ubiquitylated, endocytosed and recycled in neurons.

The neural cell adhesion molecule NCAM plays an important role during neural development and in the adult brain. To study the intracellular trafficking of NCAM in neurons, two major isoforms, NCAM140 or NCAM180, were expressed in primary cortical neurons and in the rat B35 neuroblastoma cell line. NCAM was endocytosed and subsequently recycled to the plasma membrane, whereas only a minor fraction was degraded in lysosomes. In cortical neurons, endocytosis of NCAM was detected in the soma, neurites and growth cones in a developmentally regulated fashion. Furthermore, we found that NCAM is mono-ubiquitylated at the plasma membrane and endocytosis was significantly increased in cells overexpressing ubiquitin. Therefore, we propose that ubiquitylation represents an endocytosis signal for NCAM.

Lentivirus-mediated antagomir expression for specific inhibition of miRNA function.

Micro RNAs (miRNA) regulate gene expression by hybridization and recruitment of multi-protein complexes to complementary mRNA target sequences. miRNA function can transiently be antagonized by antagomirs-chemically modified oligonucleotides complementary to individual miRNAs. Here, we describe the induction of stable loss-of-function phenotypes for specific miRNAs by lentivirus-mediated antagomir expression. Lentivirally expressed antagomirs are transcribed from a H1-promoter located within the lentiviral 3'LTR and were directed against miRNAs encoded on the polycistronic miR17-92 transcript. Functional silencing of miR-18a, miR-19b and miR-20a by the corresponding antagomirs specifically relieves miRNA-mediated reporter gene repression. Inhibition of miRNA function correlates to reduction of 'miRNA' amplification by miRNA-specific quantitative RT-PCR. Furthermore, protein expression of E2F-1, a known miR-20 target, is enhanced by lentivirally expressed anti-miR-20 antagomirs in a dose-dependent manner, whereas over-expression of miR-20a reduces E2F-1 levels. Finally, combined over-expression of specific miRNAs and antagomirs reveals individual and complementary functions of miR-18a and miR-20a and demonstrates specific miRNA impact on cell proliferation in a cell culture model.

Comparison between molecularly defined and conventional therapeutics in a conditional BCR-ABL cell culture model.

Accumulating knowledge about the molecular mechanisms causing human diseases can support the development of targeted therapies such as imatinib, a BCR-ABL-specific tyrosine kinase inhibitor to treat chronic myeloid leukemia (CML). Here, we use lentivirus-mediated RNA interference (RNAi) targeting BCR-ABL and the downstream signaling molecules SHP2, STAT5, and Gab2 to compare the efficacy and specificity of molecularly defined therapeutics with that of conventional cytotoxic drugs (cytarabine, doxorubicin, etoposide) in a conditional BCR-ABL cell culture model. IC(50) values were determined for each drug in TonB cells cultured either with interleukin-3 (IL-3) or BCR-ABL, and molecularly defined therapies were studied using lentivirally expressed shRNAs. We demonstrate that conventional anti-leukemic drugs have small or no differential effects under different cell culture conditions, whereas both imatinib and specific RNAi significantly inhibit proliferation of TonB cells in the presence of BCR-ABL but not IL-3. To study molecularly defined combination therapy, we evaluated either imatinib in TonB cells with target-specific RNAi or we used lentiviral vectors to induce combinatorial RNAi through simultaneous expression of two shRNAs. These combination therapies result in increased efficacy without loss in specificity. Interestingly, combinatorial RNAi can specifically deplete TonB cell cultures in the presence of BCR-ABL, even without targeting the oncogene itself. This model provides a tool to evaluate potential therapeutic targets and to quantify efficacy and specificity preclinically of new combination therapies in BCR-ABL-positive cells.

The graded levator hinge procedure for the correction of upper eyelid retraction (an American Ophthalmological Society thesis).

PURPOSE: Many surgical techniques have been developed to address eyelid retraction with varying results. Identifying and evaluating the anatomical and pathophysiological factors involved will assist in its surgical treatment. This prospective study evaluated the graded levator hinge procedure, in combination with a Müllerectomy and/or lateral canthoplasty when indicated, in an attempt to precisely and selectively target the pathophysiology responsible for the various causes of eyelid retraction in only one surgical session. METHODS: This is a clinical, prospective study of patients with moderate to severe eyelid retraction due to various causes who underwent the graded levator hinge procedure, in combination with a Müllerectomy and/or lateral canthoplasty when indicated. The exact amount of hinging of the levator aponeurosis, and combination with a Müllerectomy and/or lateral canthoplasty, was determined by the clinical operative findings with active cooperation from the conscious patient. RESULTS: Thirty-two consecutive patients (48 eyelids) with varying degrees of upper eyelid retraction underwent the graded levator hinge procedure in combination with a Müllerectomy and or lateral canthoplasty when indicated. The mean (+/- standard deviation) preoperative palpebral vertical fissure height was 12.4 mm (+/- 0.45 mm), and the mean postoperative palpebral fissure height was 9.0 mm (+/-0.20 mm). The mean preoperative asymmetry in the palpebral fissure height was 2.41 (+/- 0.29) mm, and the mean postoperative asymmetry was 0.59 mm (+/- 0.09), and this difference was statistically significant (P <.001). The mean reduction in the palpebral fissure height was 4.6 mm (+/- 0.29 mm) (range, 1-10 mm). The graded levator hinge procedure in combination with a Müllerectomy and or lateral canthoplasty when indicated, led to a statistically significant (P <.001) reduction in mean palpebral fissure height for all patients, the bilateral subset of patients, the unilateral subset of patients, and the thyroid-related orbitopathy subgroup. The graded levator hinge procedure in combination with a Müllerectomy and/or lateral canthoplasty when indicated led to a statistically significant reduction in palpebral fissure height, asymmetry between the eyes in the total set of patients, the unilateral set of patients, and the thyroid-related orbitopathy subset, but not in the bilaterally operated subset of patients, which were already relatively symmetric preoperatively. Postoperatively 90.6 % of all eyelids were within 1 mm of the desired postoperative level (25% were equal, 68.8% were within 0.5 mm, and 6.2% greater than 1 mm from the desired level). CONCLUSIONS: The graded levator hinge procedure, alone or in combination with a Müllerectomy and/or lateral canthoplasty, is a safe and highly effective surgical approach for the treatment of various causes of upper eyelid retraction. Through consideration of the various anatomical and pathophysiological causes of eyelid retraction, excellent functional and cosmetic results are achieved with a graded procedure tapered to the needs of each individual.