HPV16 E6/E7-mediated regulation of PiwiL1 expression induces tumorigenesis in cervical cancer cells.

PURPOSE: PiwiL1 has been reported to be over-expressed in many cancers. However, the molecular mechanism by which these proteins contribute to tumorigenesis and their regulation in cancer cells is still unclear. We intend to understand the role of PiwiL1 in tumorigenesis and also its regulation in cervical cells. METHODS: We studied the effect of loss of PiwiL1 function on tumor properties of cervical cancer cells in vitro and in vivo. Also we have looked into the effect of PiwiL1 overexpression in the malignant transformation of normal cells both in vitro and in vivo. Further RNA-seq and RIP-seq analyses were done to get insight of the direct and indirect targets of PiwiL1 in the cervical cancer cells. RESULTS: Here, we report that PiwiL1 is not only over-expressed, but also play a major role in tumor induction and progression. Abolition of PiwiL1 in CaSki cells led to a decrease in the tumor-associated properties, whereas, its upregulation conferred malignant transformation of normal HaCaT cells. Our study delineates a new link between HPV oncogenes, E6 and E7 with PiwiL1. p53 and E2F1 directly bind and differentially regulate PiwiL1 promoter in a context-dependant manner. Further, RNA-seq together with RIP-RNA-seq suggested a strong and direct role for PiwiL1 in promoting metastasis in cervical cancer cells. CONCLUSION: Our study demonstrates that PiwiL1 act as an oncogene in cervical cancer by inducing tumor-associated properties and EMT pathway. The finding that HPV oncogenes, E6/E7 can positively regulate PiwiL1 suggests a possible mechanism behind HPV-mediated tumorigenesis in cervical cancer.

The centrosomal recruitment of γ-tubulin and its microtubule nucleation activity is α-fodrin guided.

The regulation and recruitment of γ-TuRCs, the prime nucleator of microtubules, to the centrosome are still thrust areas of research. The interaction of fodrin, a sub-plasmalemmal cytoskeletal protein, with γ-tubulin is a new area of interest. To understand the cellular significance of this interaction, we show that depletion of α-fodrin brings in a significant reduction of γ-tubulin in neural cell centrosomes making it functionally under-efficient. This causes a loss of nucleation ability that cannot efficiently form microtubules in interphase cells and astral microtubules in mitosis. Fluorescence Recovery after Photobleaching (FRAP) experiment implies that α-fodrin is important in the recruitment of γ-tubulin to the centrosome resulting in the aforementioned effects. Further, our experiments indicate that the interaction of α-fodrin with certain pericentriolar matrix proteins such as Pericentrin and CDK5RAP2 are crucial for the recruitment of γ-tubulin to the centrosome. Earlier we reported that α-fodrin limits the nucleation potential of γ-TuRC. In that context, this study suggests that α-fodrin is a γ-tubulin recruiting protein to the centrosome thus preventing cytoplasmic microtubule nucleation and thereby compartmentalizing the process to the centrosome for maximum efficiency. Summary statementα-fodrin is a γ-tubulin interacting protein that controls the process of γ-tubulin recruitment to the centrosome and thereby regulates the microtubule nucleation capacity spatially and temporally.

HES1 promoter activation dynamics reveal the plasticity, stemness and heterogeneity in neuroblastoma cancer stem cells.

Notch signaling and its downstream gene target HES1 play a critical role in regulating and maintaining cancer stem cells (CSCs), similar to as they do during embryonic development. Here, we report a unique subclass of Notch-independent Hes-1 (NIHes-1)-expressing CSCs in neuroblastoma. These CSCs maintain sustained HES1 expression by activation of HES1 promoter region upstream of classical CBF-1 binding sites, thereby completely bypassing Notch receptor-mediated activation. These stem cells have self-renewal ability and potential to generate tumors. Interestingly, we observed that NIHes-1 CSCs could transition to Notch-dependent Hes-1-expressing (NDHes-1) CSCs where HES1 is expressed by Notch receptor-mediated promoter activation. We observed that NDHes-1-expressing CSCs also had the potential to transition to NIHes-1 CSCs and during this coordinated bidirectional transition, both CSCs gave rise to the majority of the bulk cancer cells, which had an inactive HES1 promoter (PIHes-1). A few of these PIHes-1 cells were capable of reverting into a CSC state. These findings explain the existence of a heterogenic mode of HES1 promoter activation within the IMR-32 neuroblastoma cell line and the potential to switch between them. This article has an associated First Person interview with the first authors of the paper.

Supramolecular Hydrogel Based Post-Surgical Implant System for Hydrophobic Drug Delivery Against Glioma Recurrence.

Purpose: The brain, protected by the cranium externally and the blood-brain barrier (BBB) internally, poses challenges in chemotherapy of aggressive brain tumors. Maximal tumor resection followed by radiation and chemotherapy is the standard treatment protocol; however, a substantial number of patients suffer from recurrence. Systemic circulation of drugs causes myelodysplasia and other side effects. To address these caveats, we report facile synthesis of a polyester-based supramolecular hydrogel as a brain biocompatible implant for in situ delivery of hydrophobic drugs. Methods: Polycaprolactone-diol (PCL) was linked to polyethyleneglycol-diacid (PEG) via an ester bond. In silico modeling indicated micelle-based aggregation of PCL-PEG co-polymer to form a supramolecular hydrogel. Brain biocompatibility was checked in Sprague Dawley rat brain cortex with MRI, motor function test, and histology. Model hydrophobic drugs carmustine and curcumin entrapment propelled glioma cells into apoptosis-based death evaluated by in vitro cytotoxicity assays and Western blot. In vivo post-surgical xenograft glioma model was developed in NOD-SCID mice and evaluated for efficacy to restrict aggressive regrowth of tumors. Results: 20% (w/v) PCL-PEG forms a soft hydrogel that can cover the uneven and large surface area of a tumor resection cavity and maintain brain density. The PCL-PEG hydrogel was biocompatible, and well-tolerated upon implantation in rat brain cortex, for a study period of 12 weeks. We report for the first time the combination of carmustine and curcumin entrapped as model hydrophobic drugs, increasing their bioavailability and yielding synergistic apoptotic effect on glioma cells. Further in vivo study indicated PCL-PEG hydrogel with a dual cargo of carmustine and curcumin restricted aggressive regrowth post-resection significantly compared with control and animals with intravenous drug treatment. Conclusion: PCL-PEG soft gel-based implant is malleable compared with rigid wafers used as implants, thus providing larger surface area contact. This stable, biocompatible, supramolecular gel without external crosslinking can find wide applications by interchanging formulation of various hydrophobic drugs to ensure and increase site-specific delivery, avoiding systemic circulation.

Pax6 modulates intra-retinal axon guidance and fasciculation of retinal ganglion cells during retinogenesis.

Intra-retinal axon guidance involves a coordinated expression of transcription factors, axon guidance genes, and secretory molecules within the retina. Pax6, the master regulator gene, has a spatio-temporal expression typically restricted till neurogenesis and fate-specification. However, our observation of persistent expression of Pax6 in mature RGCs led us to hypothesize that Pax6 could play a major role in axon guidance after fate specification. Here, we found significant alteration in intra-retinal axon guidance and fasciculation upon knocking out of Pax6 in E15.5 retina. Through unbiased transcriptome profiling between Pax6fl/fl and Pax6-/- retinas, we revealed the mechanistic insight of its role in axon guidance. Our results showed a significant increase in the expression of extracellular matrix molecules and decreased expression of retinal fate specification and neuron projection guidance molecules. Additionally, we found that EphB1 and Sema5B are directly regulated by Pax6 owing to the guidance defects and improper fasciculation of axons. We conclude that Pax6 expression post fate specification of RGCs is necessary for regulating the expression of axon guidance genes and most importantly for maintaining a conducive ECM through which the nascent axons get guided and fasciculate to reach the optic disc.

Drug treatment patterns and work productivity in chronic kidney disease patients with anemia in China: cross sectional analysis of real-world data

Background@#We explored the association of anemia severity in patients with chronic kidney disease (CKD) and anemia treatment with work productivity in China. @*Methods@#Cross-sectional survey data from Chinese physicians and their CKD patients were collected in 2015. Physicians recorded demographics, disease characteristics, and treatment. Patients completed the Work Productivity and Activity Impairment questionnaire. Data were stratified by dialysis-dependence, hemoglobin (Hb) level, and anemia treatment. @*Results@#Based on data from 1,052 patients (704 non-dialysis-dependent [NDD] and 348 dialysis-dependent [DD] patients), prescribed anemia treatment differed significantly across Hb levels (P 12 g/dL, 31%, 59%, and 38% of patients, respectively, were prescribed oral iron, and 34%, 19%, and 0% of patients, respectively, were prescribed oral iron with erythropoiesis-stimulating agents (ESA). NDD patients were less likely to be prescribed any anemia treatment, and ESA specifically, than DD patients. When treated, 67% and 45% of NDD and DD patients, respectively, had Hb ≥ 10 g/dL (P < 0.001). Overall work and activity impairment differed significantly across Hb levels in NDD and DD patients, with the least impairment observed at the highest Hb level. @*Conclusion@#Approximately 40% of NDD patients and 60% of DD patients receiving anemia treatment had Hb < 10 g/dL. Compared with mild anemia patients, severe anemia patients were more likely to be treated for anemia and have impaired work productivity. Chinese CKD patients could benefit from improved anemia treatment.

Alteration in the phosphorylation status of NMDA receptor GluN2B subunit by activation of both NMDA receptor and L-type voltage gated calcium channel.

Calcium influx through N-methyl-D-aspartate receptors (NMDAR) and voltage-gated calcium channels (VGCC) play major roles in postsynaptic signaling mechanisms. NMDAR subunit GluN2B is phosphorylated at Ser1303. Phosphorylation at this site is a prominent event in cell culture systems as well as in vivo. However, the functional significance of phosphorylation at this site is not completely understood. In this study, we compared the effect of calcium signaling through NMDAR and VGCC on the phosphorylation status of GluN2B-Ser1303 in the rat in vivo. VGCC was activated by intraperitoneal (IP) injection of the activator, BayK8644 and NMDAR was activated by intracerebroventricular (ICV) injection of NMDA in separate experimental groups. We found that the level of phospho-GluN2B-Ser1303 in the cortex and in the hippocampus increased in response to activation of either channel. The effects could be prevented by prior ICV administration of the specific blockers of these channels such as MK-801 for NMDAR and nifedipine for VGCC. The effect was also blocked by pretreatment with ICV administration of KN-93 indicating that it is mediated through CaM kinase. Both during NMDAR activation and VGCC activation, cell survival associated signals such as phospho-AKT and phospho-CREB showed decrease, consistent with activation of cell death pathways during these treatments. We conclude that under in vivo conditions, calcium influx through either NMDAR or VGCC activates CaM kinase, which in turn phosphorylates GluN2B-Ser1303.

Pneumomediastinum and pneumopericardium following blunt thoracic trauma: much ado about nothing?

PURPOSE: Pneumomediastinum is the hallmark of intrathoracic aerodigestive trauma, but rare following blunt injury. AIM: review of blunt thoracic trauma (BTC) for the incidence and outcome of patients with pneumomediastinum or pneumopericardium (PM/PC) on Computerised Tomographic scanning. METHODS: Admissions to the level I trauma ICU at IALCH, Durban, ZA following BTC from April 2007 to March 2014. Patients with Chest-CT-scan were analysed. Variables included age, sex, mechanism of injury, and Injury Severity Score (ISS). Specific injury patterns: isolated thoracic trauma, flail chest, bilateral injury and presence of haemothorax or pneumothorax were analysed. RESULTS: Three hundred and eighty-nine patients were included. Males (70.9%) accounted for the majority of patients. The median Injury Severity Score was 32 (IQR 24-41). Motor vehicle collisions accounted for 94% of injury mechanisms. Twenty-three (5.9%) were identified with pneumomediastinum, 6 (1.5%) with both pneumomediastinum and pneumopericardium, and 1 (0.2%) with isolated pneumopericardium. No patient required surgery for thoracic trauma. Increasing age (p < 0.001) and a flail chest (p = 0.005) were significant associations. The mortality rate was almost identical in those with or without air within the mediastinum. No patient died from a missed mediastinal aero-digestive injury. CONCLUSION: The presence of PM/PC following BTC is incidental and benign. Increased injury severity with a flail chest is associated with a significant increase in the presence of free gas within the mediastinum. In the absence of complications, no obvious injury to the intrathoracic aero-digestive tract on CT scanning, and no difference in mortality, a conservative management policy is warranted.

Intraocular Injection of ES Cell-Derived Neural Progenitors Improve Visual Function in Retinal Ganglion Cell-Depleted Mouse Models.

Retinal ganglion cell (RGC) transplantation is a promising strategy to restore visual function resulting from irreversible RGC degeneration occurring in glaucoma or inherited optic neuropathies. We previously demonstrated FGF2 induced differentiation of mouse embryonic stem cells (ESC) to RGC lineage, capable of retinal ganglion cell layer (GCL) integration upon transplantation. Here, we evaluated possible improvement of visual function by transplantation of ES cell derived neural progenitors in RGC depleted glaucoma mice models. ESC derived neural progenitors (ES-NP) were transplanted into N-Methyl-D-Aspartate (NMDA) injected, RGC-ablated mouse models and a pre-clinical glaucoma mouse model (DBA/2J) having sustained higher intra ocular pressure (IOP). Visual acuity and functional integration was evaluated by behavioral experiments and immunohistochemistry, respectively. GFP-expressing ES-NPs transplanted in NMDA-injected RGC-depleted mice differentiated into RGC lineage and possibly integrating into GCL. An improvement in visual acuity was observed after 2 months of transplantation, when compared to the pre-transplantation values. Expression of c-Fos in the transplanted cells, upon light induction, further suggests functional integration into the host retinal circuitry. However, the transplanted cells did not send axonal projections into optic nerve. Transplantation experiments in DBA/2J mouse showed no significant improvement in visual functions, possibly due to both host and transplanted retinal cell death which could be due to an inherent high IOP. We showed that, ES NPs transplanted into the retina of RGC-ablated mouse models could survive, differentiate to RGC lineage, and possibly integrate into GCL to improve visual function. However, for the survival of transplanted cells in glaucoma, strategies to control the IOP are warranted.

Wnt5a is a crucial regulator of neurogenesis during cerebellum development.

The role of Wnt5a has been extensively explored in various aspects of development but its role in cerebellar development remains elusive. Here, for the first time we unravel the expression pattern and functional significance of Wnt5a in cerebellar development using Wnt5a-/- and Nestin-Cre mediated conditional knockout mouse models. We demonstrate that loss of Wnt5a results in cerebellar hypoplasia and depletion of GABAergic and glutamatergic neurons. Besides, Purkinje cells of the mutants displayed stunted, poorly branched dendritic arbors. Furthermore, we show that the overall reduction is due to decreased radial glial and granule neuron progenitor cell proliferation. At molecular level we provide evidence for non-canonical mode of action of Wnt5a and its regulation over genes associated with progenitor proliferation. Altogether our findings imply that Wnt5a signaling is a crucial regulator of cerebellar development and would aid in better understanding of cerebellar disease pathogenesis caused due to deregulation of Wnt signaling.

Pleiotropic Hes-1 Concomitant with its Differential Activation Mediates Neural Stem Cell Maintenance and Radial Glial Propensity in Developing Neocortex.

Notch signaling pathway and its downstream effector Hes-1 are well known for their role in cortical neurogenesis. Despite the canonical activation of Hes-1 in developing neocortex, recent advances have laid considerable emphasis on Notch/CBF1-independent Hes-1 (NIHes-1) expression with poor understanding of its existence and functional significance. Here, using reporter systems and in utero electroporation, we could qualitatively unravel the existence of NIHes-1 expressing neural stem cells from the cohort of dependent progenitors throughout the mouse neocortical development. Though Hes-1 expression is maintained in neural progenitor territory at all times, a simple shift from Notch-independent to -dependent state makes it pleiotropic as the former maintains the neural stem cells in a non-dividing/slow-dividing state, whereas the latter is very much required for maintenance and proliferation of radial glial cells. Therefore, our results provide an additional complexity in neural progenitor heterogeneity regarding differential Hes-1 expression in the germinal zone during neo-cortical development.

Regulation of Tlx3 by Pax6 is required for the restricted expression of Chrnα3 in Cerebellar Granule Neuron progenitors during development.

Homeobox gene Tlx3 is known to promote glutamatergic differentiation and is expressed in post-mitotic neurons of CNS. Contrary to this here, we discovered that Tlx3 is expressed in the proliferating progenitors of the external granule layer in the cerebellum, and examined factors that regulate this expression. Using Pax6(-/-)Sey mouse model and molecular interaction studies we demonstrate Pax6 is a key activator of Tlx3 specifically in cerebellum, and induces its expression starting at embryonic day (E)15. By Postnatal day (PN)7, Tlx3 is expressed in a highly restricted manner in the cerebellar granule neurons of the posterior cerebellar lobes, where it is required for the restricted expression of nicotinic cholinergic receptor-α3 subunit (Chrnα3) and other genes involved in formation of synaptic connections and neuronal migration. These results demonstrate a novel role for Tlx3 and indicate that Pax6-Tlx3 expression and interaction is part of a region specific regulatory network in cerebellum and its deregulation during development could possibly lead to Autistic spectral disorders (ASD).

Congenital lobar emphysema: diagnostic and therapeutic challenges.

Congenital lobar emphysema (CLE), a rare condition that usually presents in the neonatal period, can be a diagnostic and therapeutic challenge for the treating clinician. If unrecognised, it is a significant risk at the time of anaesthetic induction. We describe a case of CLE in a 3-month-old boy who was initially treated for suspected aspiration pneumonia at the referring hospital. We highlight the importance of careful consideration of common childhood respiratory illness as well as pneumothorax in the differential diagnosis, and the significance of appropriate preoperative anaesthetic management. We also emphasise the importance of acknowledging a mother's concerns when taking a paediatric history.

Hes1: the maestro in neurogenesis.

The process of neurogenesis is well orchestrated by the harmony of multiple cues in a spatiotemporal manner. In this review, we focus on how a dynamic gene, Hes1, is involved in neurogenesis with the view of its regulation and functional implications. Initially, we have reviewed the immense functional significance drawn by this maestro during neural development in a context-dependent manner. How this indispensable role of Hes1 in conferring the competency for neural differentiation partly relies on the direct/indirect mode of repression mediated by very specific structural and functional arms of this protein has also been outlined here. We also review the detailed molecular mechanisms behind the well-tuned oscillatory versus sustained expression of this antineurogenic bHLH repressor, which indeed makes it a master gene to implement the elusive task of neural progenitor propensity. Apart from the functional aspects of Hes1, we also discuss the molecular insights into the endogenous regulatory machinery that regulates its expression. Though Hes1 is a classical target of the Notch signaling pathway, we discuss here its differential expression at the molecular, cellular, and/or regional level. Moreover, we describe how its expression is fine-tuned by all possible ways of gene regulation such as epigenetic, transcriptional, post-transcriptional, post-translational, and environmental factors during vertebrate neurogenesis.

Molecular mechanisms in H2O2-induced increase in AT1 receptor gene expression in cardiac fibroblasts: A role for endogenously generated Angiotensin II.

The AT1 receptor (AT1R) mediates the manifold actions of angiotensin II in the cardiovascular system. This study probed the molecular mechanisms that link altered redox status to AT1R expression in cardiac fibroblasts. Real-time PCR and western blot analysis showed that H2O2 enhances AT1R mRNA and protein expression via NADPH oxidase-dependent reactive oxygen species induction. Activation of NF-κB and AP-1, demonstrated by electrophoretic mobility shift assay, abolition of AT1R expression by their inhibitors, Bay-11-7085 and SR11302, respectively, and luciferase and chromatin immunoprecipitation assays confirmed transcriptional control of AT1R by NF-κB and AP-1 in H2O2-treated cells. Further, inhibition of ERK1/2, p38 MAPK and c-Jun N-terminal kinase (JNK) using chemical inhibitors or by RNA interference attenuated AT1R expression. Inhibition of the MAPKs showed that while ERK1/2 and p38 MAPK suffice for NF-κB activation, all three kinases are required for AP-1 activation. H2O2 also increased collagen type I mRNA and protein expression. Interestingly, the AT1R antagonist, candesartan, attenuated H2O2-stimulated AT1R and collagen mRNA and protein expression, suggesting that H2O2 up-regulates AT1R and collagen expression via local Angiotensin II generation, which was confirmed by real-time PCR and ELISA. To conclude, oxidative stress enhances AT1R gene expression in cardiac fibroblasts by a complex mechanism involving the redox-sensitive transcription factors NF-κB and AP-1 that are activated by the co-ordinated action of ERK1/2, p38 MAPK and JNK. Importantly, by causally linking oxidative stress to Angiotensin II and AT1R up-regulation in cardiac fibroblasts, this study offers a novel perspective on the pathogenesis of cardiovascular diseases associated with oxidative stress.

Empirical antimicrobial therapy for probable v. directed therapy for possible ventilator-associated pneumonia in critically injured patients.

BACKGROUND: Ventilator-associated pneumonia (VAP) has recently been classified as possible or probable. Although direct attributable mortality has been difficult to prove, delay in instituting appropriate therapy has been reported to increase morbidity and mortality. Recent literature suggests that in possible VAP, instituting directed therapy while awaiting microbiological culture does not prejudice outcome compared with best-guess empirical therapy. OBJECTIVES: To ascertain outcomes of directed v. empirical therapy in possible and probable VAP, respectively. METHODS: Endotracheal aspirates were obtained from patients with suspected VAP. Those considered to have possible VAP were given directed therapy following culture results, whereas patients with more convincing evidence of VAP were classed as having probable VAP and commenced on empirical antimicrobials based on microbiological surveillance. RESULTS: Pneumonia was suspected in 106 (36.8%) of 288 patients admitted during January - December 2014. Of these, 13 did not fulfil the criteria for VAP. Of the remaining 93 (32.2%), 31 (33.3%) were considered to have probable and 62 (66.7%) possible VAP. The former were commenced on empirical antimicrobials, with 28 (90.3%) receiving appropriate therapy. Of those with possible VAP, 34 (54.8%) were given directed therapy and in 28 (45.2%) no antimicrobials were prescribed. Of the latter, 24 recovered without antimicrobials and 4 died, 3 from severe traumatic brain injury and 1 due to overwhelming intra-abdominal sepsis. No death was directly attributable to failure to treat VAP. No significant difference in mortality was found between the 34 patients with possible VAP who were commenced on directed therapy and the 31 with probable VAP who were commenced on empirical antimicrobials (p=0.75). CONCLUSIONS: Delaying antimicrobial therapy for VAP where clinical doubt exists does not adversely affect outcome. Furthermore, this policy limits the use of antimicrobials in patients with possible VAP following improvement in their clinical condition despite no therapy.

Molecular basis and functional significance of Angiotensin II-induced increase in Discoidin Domain Receptor 2 gene expression in cardiac fibroblasts.

Delineation of mechanisms underlying the regulation of fibrosis-related genes in the heart is an important clinical goal as cardiac fibrosis is a major cause of myocardial dysfunction. This study probed the regulation of Discoidin Domain Receptor 2 (DDR2) gene expression and the regulatory links between Angiotensin II, DDR2 and collagen in Angiotensin II-stimulated cardiac fibroblasts. Real-time PCR and western blot analyses showed that Angiotensin II enhances DDR2 mRNA and protein expression in rat cardiac fibroblasts via NADPH oxidase-dependent reactive oxygen species induction. NF-κB activation, demonstrated by gel shift assay, abolition of DDR2 expression upon NF-κB inhibition, and luciferase and chromatin immunoprecipitation assays confirmed transcriptional control of DDR2 by NF-κB in Angiotensin II-treated cells. Inhibitors of Phospholipase C and Protein kinase C prevented Angiotensin II-dependent p38 MAPK phosphorylation that in turn blocked NF-κB activation. Angiotensin II also enhanced collagen gene expression. Importantly, the stimulatory effects of Angiotensin II on DDR2 and collagen were inter-dependent as siRNA-mediated silencing of one abolished the other. Angiotensin II promoted ERK1/2 phosphorylation whose inhibition attenuated Angiotensin II-stimulation of collagen but not DDR2. Furthermore, DDR2 knockdown prevented Angiotensin II-induced ERK1/2 phosphorylation, indicating that DDR2-dependent ERK1/2 activation enhances collagen expression in cells exposed to Angiotensin II. DDR2 knockdown was also associated with compromised wound healing response to Angiotensin II. To conclude, Angiotensin II promotes NF-κB activation that up-regulates DDR2 transcription. A reciprocal regulatory relationship between DDR2 and collagen, involving cross-talk between the GPCR and RTK pathways, is central to Angiotensin II-induced increase in collagen expression in cardiac fibroblasts.

Treatment of hypertrophic distal tibia nonunion and early malunion with callus distraction.

BACKGROUND: Hypertrophic nonunions and early malunions of pilon and distal tibia fractures result in complex, challenging to treat deformities. Callus distraction histiogenesis is an option for their management, allowing for the simultaneous correction of multiplanar deformity and limb length discrepancy. METHODS: A single-surgeon, retrospective case series was performed. Eight patients (6 males and 2 females) who were treated with callus distraction from 1991 to 2011 were reviewed. Six of 8 patients had varus deformities (range, 8-19 degrees) and 2 patients had valgus deformities (both 16 degrees) of the distal tibia metaphysis. Six of 8 had apex anterior deformities (range, 2-21 degrees) and 2 had apex posterior deformity (range, 9-20 degrees). An Ilizarov fixator or Taylor Spatial Frame (Smith & Nephew, Memphis, TN) was used to distract the nonunion or early malunion to correct alignment and shortening. Preoperative and postoperative radiographic outcomes, limb alignment, and ankle-hindfoot scores were reviewed. RESULTS: Union was achieved in all patients at a mean of 5.8 months (range, 4.1-7.6 months). The 3 patients treated with an Ilizarov-type fixator had deformity correction to within 5 degrees of neutral in 1 plane and to within 10 degrees in the other plane. All 5 patients treated with a Taylor Spatial Frame had correction to within 5 degrees of neutral alignment in both coronal and sagittal planes. There were 2 complications requiring reoperation and 1 persistent limb length discrepancy (2 cm) after treatment. Median AOFAS ankle-hindfoot score was 82.5 (range, 53-90) at an average follow-up of 30.4 months (range, 8-92). CONCLUSIONS: Callus distraction histiogenesis was a minimally invasive technique that can successfully treat patients with hypertrophic nonunion and early malunion of the distal tibia. We believe the application of a computer-assisted 6-axis frame to correct the deformity improved the correction of these multiplanar deformities. LEVEL OF EVIDENCE: Level IV, case series.

Developmental wave of Brn3b expression leading to RGC fate specification is synergistically maintained by miR-23a and miR-374.

Differential regulation of Brn3b is essential for the Retinal Ganglion Cell (RGC) development in the two phases of retinal histogenesis. This biphasic Brn3b regulation is required first, during early retinal histogenesis for RGC fate specification and secondly, during late histogenesis, where Brn3b is needed for RGC axon guidance and survival. Here, we have looked into how the regulation of Brn3b at these two stages happens. We identified two miRNAs, miR-23a and miR-374, as regulators of Brn3b expression, during the early stage of RGC development. Temporal expression pattern of miR-23a during E10-19, PN1-7, and adult retina revealed an inverse relation with Brn3b expression. Though miR-374 did not show such a pattern, its co-expression with miR-23a evidently inhibited Brn3b. We further substantiated these findings by ex vivo overexpression of these miRNAs in E14 mice retina and found that miR-23a and miR-374 together brings about a change in Brn3b expression pattern in ganglion cell layer (GCL) of the developing retina. From our results, it appears that the combined expression of these miRNAs could be regulating the timing of the wave of Brn3b expression required for early ganglion cell fate specification and later for its survival and maturation into RGCs. Taken together, here we provide convincing evidences for the existence of a co-ordinated mechanism by miRNAs to down regulate Brn3b that will ultimately regulate the development of RGCs from their precursors.

Morphology and electrostatics play active role in neuronal differentiation processes on flexible conducting substrates.

This commentary discusses and summarizes the key highlights of our recently reported work entitled "Neuronal Differentiation of Embryonic Stem Cell Derived Neuronal Progenitors Can Be Regulated by Stretchable Conducting Polymers." The prospect of controlling the mechanical-rigidity and the surface conductance properties offers a unique combination for tailoring the growth and differentiation of neuronal cells. We emphasize the utility of transparent elastomeric substrates with coatings of electrically conducting polymer to realize the desired substrate-characteristics for cellular development processes. Our study showed that neuronal differentiation from ES cells is highly influenced by the specific substrates on which they are growing. Thus, our results provide a better strategy for regulated neuronal differentiation by using such functional conducting surfaces.