Author Correction: Improving Stem Cell Delivery to the Trabecular Meshwork Using Magnetic Nanoparticles.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Improving Stem Cell Delivery to the Trabecular Meshwork Using Magnetic Nanoparticles.

Glaucoma is a major cause of blindness and is frequently associated with elevated intraocular pressure. The trabecular meshwork (TM), the tissue that primarily regulates intraocular pressure, is known to have reduced cellularity in glaucoma. Thus, stem cells, if properly delivered to the TM, may offer a novel therapeutic option for intraocular pressure control in glaucoma patients. For this purpose, targeted delivery of stem cells to the TM is desired. Here, we used magnetic nanoparticles (Prussian blue nanocubes [PBNCs]) to label mesenchymal stem cells and to magnetically steer them to the TM following injection into the eye's anterior chamber. PBNC-labeled stem cells showed increased delivery to the TM vs. unlabeled cells after only 15-minute exposure to a magnetic field. Further, PBNC-labeled mesenchymal stem cells could be delivered to the entire circumference of the TM, which was not possible without magnetic steering. PBNCs did not affect mesenchymal stem cell viability or multipotency. We conclude that this labeling approach allows for targeted, relatively high-efficiency delivery of stem cells to the TM in clinically translatable time-scales, which are necessary steps towards regenerative medicine therapies for control of ocular hypertension in glaucoma patients.

ECM1 regulates tumor metastasis and CSC-like property through stabilization of ß-catenin.

Extracellular Matrix Protein 1 (ECM1) is a marker for tumorigenesis and is correlated with invasiveness and poor prognosis in various types of cancer. However, the functional role of ECM1 in cancer metastasis is unclear. Here, we detected high ECM1 level in breast cancer patient sera that was associated with recurrence of tumor. The modulation of ECM1 expression affected not only cell migration and invasion, but also sphere-forming ability and drug resistance in breast cancer cell lines. In addition, ECM1 regulated the gene expression associated with the epithelial to mesenchymal transition (EMT) progression and cancer stem cell (CSC) maintenance. Interestingly, ECM1 increased ß-catenin expression at the post-translational level through induction of MUC1, which was physically associated with ß-catenin. Indeed, the association between ß-catenin and the MUC1 cytoplasmic tail was increased by ECM1. Furthermore, forced expression of ß-catenin altered the gene expression that potentiated EMT progression and CSC phenotype maintenance in the cells. These data provide evidence that ECM1 has an important role in cancer metastasis through ß-catenin stabilization.

PKCδ maintains phenotypes of tumor initiating cells through cytokine-mediated autocrine loop with positive feedback.

The existence of tumor initiating cells (TICs) has been emerged as a good therapeutic target for treatment of glioblastoma that is the most aggressive brain tumor with poor prognosis. However, the molecular mechanisms that regulate the phenotypes of TICs still remain obscure. In this study, we found that PKCδ, among PKC isoforms, is preferentially activated in TICs and acts as a critical regulator for the maintenance of TICs in glioblastoma. By modulating the expression levels or activity of PKCδ, we demonstrated that PKCδ promotes self-renewal and tumorigenic potentials of TICs. Importantly, we found that the activation of PKCδ persists in TICs through an autocrine loop with positive feedback that was driven by PKCδ/STAT3/IL-23/JAK signaling axis. Moreover, for phenotypes of TICs, we showed that PKCδ activates AKT signaling component by phosphorylation specifically on Ser473. Taken together, we proposed that TICs regulate their own population in glioblastoma through an autocrine loop with positive feedback that is driven by PKCδ-dependent secretion of cytokines.

Novel signaling axis for ROS generation during K-Ras-induced cellular transformation.

Reactive oxygen species (ROS) are well known to be involved in oncogene-mediated cellular transformation. However, the regulatory mechanisms underlying ROS generation in oncogene-transformed cells are unclear. In the present study, we found that oncogenic K-Ras induces ROS generation through activation of NADPH oxidase 1 (NOX1), which is a critical regulator for the K-Ras-induced cellular transformation. NOX1 was activated by K-Ras-dependent translocation of p47(phox), a subunit of NOX1 to plasma membrane. Of note, PKCδ, when it was activated by PDPK1, directly bound to the SH3-N domain of p47(phox) and catalyzed the phosphorylation on Ser348 and Ser473 residues of p47(phox) C-terminal in a K-Ras-dependent manner, finally leading to its membrane translocation. Notably, oncogenic K-Ras activated all MAPKs (JNK, ERK and p38); however, only p38 was involved in p47(phox)-NOX1-dependent ROS generation and consequent transformation. Importantly, K-Ras-induced activation of p38 led to an activation of PDPK1, which then signals through PKCδ, p47(phox) and NOX1. In agreement with the mechanism, inhibition of p38, PDPK1, PKCδ, p47(phox) or NOX1 effectively blocked K-Ras-induced ROS generation, anchorage-independent colony formation and tumor formation. Taken together, our findings demonstrated that oncogenic K-Ras activates the signaling cascade p38/PDPK1/PKCδ/p47(phox)/NOX1 for ROS generation and consequent malignant cellular transformation.

Claudin-1 induces epithelial-mesenchymal transition through activation of the c-Abl-ERK signaling pathway in human liver cells.

Claudins (CLDNs) are a family of integral membrane proteins central to the formation of tight junctions, structures that are involved in paracellular transport and cellular growth and differentiation, and are critical for the maintenance of cellular polarity. Recent studies have provided evidence that CLDNs are aberrantly expressed in diverse types of human cancers, including hepatocellular carcinomas (HCCs). However, little is known about how CLDN expression is involved in cancer progression. In this study, we show that CLDN1 has a causal role in the epithelial-mesenchymal transition (EMT) in human liver cells, and that the c-Abl-Ras-Raf-1-ERK1/2 signaling axis is critical for the induction of malignant progression by CLDN1. Overexpression of CLDN1 induced expression of the EMT-regulating transcription factors Slug and Zeb1, and thereby led to repression of E-cadherin, ß-catenin expression, enhanced expression of N-cadherin and Vimentin, a loss of cell adhesion, and increased cell motility in normal liver cells and HCC cells. In line with these findings, inhibition of either c-Abl or ERK clearly attenuated CLDN1-induced EMT, as evidenced by a reversal of N-cadherin and E-cadherin expression patterns, and restored normal motility. Collectively, these results indicate that CLDN1 is necessary for the induction of EMT in human liver cells, and that activation of the c-Abl-Ras-Raf-1-ERK1/2 signaling pathway is required for CLDN1-induced acquisition of the malignant phenotype. The present observations suggest that CLDN1 could be exploited as a biomarker for liver cancer metastasis and might provide a pivotal point for therapeutic intervention in HCC.

c-Jun N-terminal kinase has a pivotal role in the maintenance of self-renewal and tumorigenicity in glioma stem-like cells.

Uncovering the mechanisms that govern the maintenance of stem-like cancer cells is critical for developing therapeutic strategies for targeting these cells. Constitutive activation of c-Jun N-terminal kinase (JNK) has been reported in gliomas and correlates with histological grade. Here, we found that JNK signaling is crucial for the maintenance of 'stemness' in glioma cells. Sphere-cultured glioma cells showed more phosphorylation of JNK compared with serum-containing monolayer cultures. Importantly, blockade of JNK signaling with SP600125 or small interfering RNAs targeting JNK1 or JNK2 significantly reduced the CD133(+)/Nestin(+) population and suppressed sphere formation, colony formation in soft agar, and expression of stem cell markers in sphere-cultured glioma cells. Intriguingly, sphere-cultured glioma cells exhibited enhanced expression of Notch-2, but not Notch-1, -3 or -4, and JNK inhibition almost completely abrogated this increase. Blocking the phosphoinoside 3-kinase (PI3K)/Akt pathway with LY294002 or si-Akt also suppressed the self-renewal of sphere-cultured glioma cells. PI3K, but not Akt, had a role as an upstream kinase in JNK1/2 activation. In addition, treatment with si-JNK greatly increased etoposide- and ionizing radiation (IR)-induced cell death in glioma spheres. Consistent with glioma cell lines, glioma stem-like cells isolated from primary patient glioma cells also had a higher activity of JNK and Notch-2 expression. Importantly, inhibition of JNK2 led to a decrease of Notch-2 expression and suppressed the CD133(+)/Nestin(+) cell population in patient-derived primary glioma cells. Finally, downregulation of JNK2 almost completely suppressed intracranial tumor formation by glioma cells in nude mice. Taken together, these data demonstrate that JNK signaling is crucial for the maintenance of self-renewal and tumorigenicity of glioma stem-like cells and drug/IR resistance, and can be considered a promising target for eliminating stem-like cancer cells in gliomas.

The Laurentian Great Lakes strain (MI03) of the viral haemorrhagic septicaemia virus is highly pathogenic for juvenile muskellunge, Esox masquinongy (Mitchill).

The Great Lakes strain of viral haemorrhagic septicaemia virus (VHSV) isolated from adult subclinical muskellunge, Esox masquinongy (Mitchill), in Lake St. Clair, MI, USA was shown to be highly pathogenic in juvenile muskellunge through intraperitoneal (i.p.) injection and waterborne challenge. Mortality began as early as 3 days after exposure in waterborne challenged fish, whereas fish infected by the i.p. route experienced the first mortality by 5 days post-infection (p.i.). The median lethal intraperitoneal injection dose (IP-LD(50)) was approximately 2.21 plaque forming units (PFU) as opposed to the median lethal immersion challenge dose (IM-LD(50)) of 1.7 x 10(4) PFU mL(-1). A high, medium and low dose of infection caused acute, subacute and chronic progression of the disease, respectively, as was evident by the cumulative mortality data. Clinical signs of disease observed in dead and moribund fish were very pale gills, dermal petechial haemorrhages along the flanks, severe nuchal haemorrhages, intramuscular haemorrhages at the fin-muscle junction and focal haemorrhaging on the caudal peduncle. Internal lesions included livers that were pale, discoloured and friable, and kidneys that were either congested or degenerative in appearance, and petechial to ecchymotic haemorrhages on the swim bladder wall. Histopathologic examination demonstrated massive haemorrhages in the swimbladder wall and muscle, severe vacuolation and multifocal necrosis of the liver, multifocal necrosis of the gills and depletion of lymphoid tissues within the spleen. Kidney tissues also exhibited a mixed pattern of degeneration that included tubular necrosis, interstitial oedema and congestion. Virus was recovered from kidney and spleen tissues through tissue culture and reverse transcriptase-polymerase chain reaction (RT-PCR).

Design and reliability of pediatric HealthQuiz: preliminary report of a comprehensive, computerized, self-administered child health assessment.

The time available for pediatric ambulatory visits is rarely sufficient to permit a truly comprehensive health assessment. We hypothesized that a reliable, computerized, self-administered questionnaire could be designed to screen for a full range of pediatric health issues and provide a comprehensive health database for pediatric patients. An age- and gender-specific pediatric questionnaire of 478 questions was formatted to elicit only a "Yes," "No," or "Not Sure" response and structured in a branched, decision-tree format. The initial draft was reviewed for content by pediatric experts in Canada and the United States and revised in accordance with their suggestions. The questionnaire was divided into two modules, Medical Peds, covering biomedical issues and Prevent Peds, covering prevention, psychosocial, educational, and safety topics. Cognitive interviews were carried out with 132 parents in pediatric ambulatory care centers in Chicago and Halifax, with use of scripted and nonscripted probe questions, to ensure comprehensibility among patients with widely varying educational levels and health knowledge. Reliability was tested in 100 parents of children aged 1 month to 12 years, through use of five different test-retest sequences. Respondents' impressions were surveyed on completion of the procedure. Following content reviews, and cognitive and reliability testing, the total bank of questions was reduced to 375. As a result of the use of branching logic, individual parents answered an average of 111 Prevent Peds and 144 Medical Peds questions. Average time required to complete the entire questionnaire was 13 minutes for Prevent Peds and 19 minutes for Med Peds. Retesting within 36 hours showed an overall 97% concordance of response pairs in the Medical Peds and Prevent Peds questionnaires. There were no statistically significant differences in test-retest reliability between different sequence formats used, (e.g., HealthQuiz followed by personal interview, or HealthQuiz vs. HealthQuiz). A few questions that frequently elicited "Not Sure" responses were eliminated. As a result, the majority of questions elicited either a "Yes" or "No" response. Pediatric HealthQuiz identified a wide spectrum of child health problems that are often overlooked in routine health visits. Parents completing Pediatric HealthQuiz indicated a high degree of satisfaction with the procedure. Most reported that they believed the information would improve their child's health care.

[Effect of ascorbic acid on 25-hydroxyvitamin D3 metabolism in the kidneys and 1,25-dihydroxyvitamin D3 reception in the small intestine mucosa of guinea pigs]. / Vliianie askorbinovoi kisloty na obmen 25-oksivitamina D3 v pochkakh i retseptsiiu 1,25-dioksivitamina D3 v slizistoi obolochke tonkogo kishechnika u morskikh svinok.

Ascorbic acid deficiency in vitamin D-supplied guinea pigs caused a moderate decrease of Ca in the blood and osseous tissue, a 1.5-fold decrease of 2.5-hydroxyvitamin D (25-OH D) in blood serum, a 2-fold decrease of the 25-OH D 1-hydroxylase activity in kidneys and a 1.6-fold increase of the 24-hydroxylase activity. The concentration of 1.25-dihydroxyvitamin D3 (1.25-(OH)2D3) nuclear receptors in small intestinal mucosa diminished by 20-30%; in this case the percentage of occupied hormone receptors reduced from 11.8 to 8.6%. The affinity of receptors for 1.25-(OH)2D3 did not change thereby (Kd = 0.25-0.26 nM; Kd2 = 0.06-0.10 nM). At the same time the value of cooperativity coefficient showed a decrease-from 1.7 to 1.4, which was accompanied by a reduction of the maximum capacity of receptors (1.2-1.5-fold). Vitamin C depletion augmented the manifestation of vitamin D deficiency in guinea pigs and impeded their correction after administration of cholecalciferol. This markedly retarded the restoration of the 25-OH D level in the blood as well as the number of occupied and unoccupied nuclear receptors for 1.25-(OH)2D3. The experimental results illustrate the effects of ascorbic acid on the vitamin D hormonal system function, which is manifested both at the level of 1.25-(OH)2D3 synthesis in the kidneys and of its receptor binding in target tissues.

Ultraviolet-visible absorption in highly transparent solids by laser calorimetry and wavelength modulation spectroscopy.

The requirements for low-loss optical materials for use on excimer lasers have stimulated the investigation of optical absorption in a variety of highly transparent materials at visible and uv wavelengths. To provide information over a wide spectral range at low absorption levels ( approximately 10(-5) cm(-1)), laser calorimetric and wavelength modulation spectroscopic techniques were used. Blending these two methods provided, for the first time, spectral information well below the usual levels of absorption measured in studies of the Urbach tail.