EYA protein complex is required for Wntless retrograde trafficking from endosomes to Golgi.

Retrograde transport of WLS (Wntless) from endosomes to trans-Golgi network (TGN) is required for efficient Wnt secretion during development. However, the molecular players connecting endosomes to TGN during WLS trafficking are limited. Here, we identified a role for Eyes Absent (EYA) proteins during retrograde trafficking of WLS to TGN in human cell lines. By using worm, fly, and zebrafish models, we found that the EYA-secretory carrier-associated membrane protein 3 (SCAMP3) axis is evolved in vertebrates. EYAs form a complex and interact with retromer on early endosomes. Retromer-bound EYA complex recruits SCAMP3 to endosomes, which is necessary for the fusion of WLS-containing endosomes to TGN. Loss of EYA complex or SCAMP3 leads to defective transport of WLS to TGN and failed Wnt secretion. EYA mutations found in patients with hearing loss form a dysfunctional EYA-retromer complex that fails to activate Wnt signaling. These findings identify the EYA complex as a component of retrograde trafficking of WLS from the endosome to TGN.

Inorganic Biomaterials Shape the Transcriptome Profile to Induce Endochondral Differentiation.

Minerals play a vital role, working synergistically with enzymes and other cofactors to regulate physiological functions including tissue healing and regeneration. The bioactive characteristics of mineral-based nanomaterials can be harnessed to facilitate in situ tissue regeneration by attracting endogenous progenitor and stem cells and subsequently directing tissue-specific differentiation. Here, cellular responses of human mesenchymal stem/stromal cells to traditional bioactive mineral-based nanomaterials, such as hydroxyapatite, whitlockite, silicon-dioxide, and the emerging synthetic 2D nanosilicates are investigated. Transcriptome sequencing is utilized to probe the cellular response and determine the significantly affected signaling pathways due to exposure to these inorganic nanomaterials. Transcriptome profiles of stem cells treated with nanosilicates reveals a stabilized skeletal progenitor state suggestive of endochondral differentiation. This observation is bolstered by enhanced deposition of matrix mineralization in nanosilicate treated stem cells compared to control or other treatments. Specifically, use of 2D nanosilicates directs osteogenic differentiation of stem cells via activation of bone morphogenetic proteins and hypoxia-inducible factor 1-alpha signaling pathway. This study provides  insight into impact of nanomaterials on cellular gene expression profile and predicts downstream effects of nanomaterial induction of endochondral differentiation.

Stiffness assisted cell-matrix remodeling trigger 3D mechanotransduction regulatory programs.

Engineered matrices provide a valuable platform to understand the impact of biophysical factors on cellular behavior such as migration, proliferation, differentiation, and tissue remodeling, through mechanotransduction. While recent studies have identified some mechanisms of 3D mechanotransduction, there is still a critical knowledge gap in comprehending the interplay between 3D confinement, ECM properties, and cellular behavior. Specifically, the role of matrix stiffness in directing cellular fate in 3D microenvironment, independent of viscoelasticity, microstructure, and ligand density remains poorly understood. To address this gap, we designed a nanoparticle crosslinker to reinforce collagen-based hydrogels without altering their chemical composition, microstructure, viscoelasticity, and density of cell-adhesion ligand and utilized it to understand cellular dynamics. This crosslinking mechanism utilizes nanoparticles as crosslink epicenter, resulting in 10-fold increase in mechanical stiffness, without other changes. Human mesenchymal stem cells (hMSCs) encapsulated in 3D responded to mechanical stiffness by displaying circular morphology on soft hydrogels (5 kPa) and elongated morphology on stiff hydrogels (30 kPa). Stiff hydrogels facilitated the production and remodeling of nascent extracellular matrix (ECM) and activated mechanotransduction cascade. These changes were driven through intracellular PI3AKT signaling, regulation of epigenetic modifiers and activation of YAP/TAZ signaling. Overall, our study introduces a unique biomaterials platform to understand cell-ECM mechanotransduction in 3D for regenerative medicine as well as disease modelling.

Bendless is essential for PINK1-Park mediated Mitofusin degradation under mitochondrial stress caused by loss of LRPPRC.

Cells under mitochondrial stress often co-opt mechanisms to maintain energy homeostasis, mitochondrial quality control and cell survival. A mechanistic understanding of such responses is crucial for further insight into mitochondrial biology and diseases. Through an unbiased genetic screen in Drosophila, we identify that mutations in lrpprc2, a homolog of the human LRPPRC gene that is linked to the French-Canadian Leigh syndrome, result in PINK1-Park activation. While the PINK1-Park pathway is well known to induce mitophagy, we show that PINK1-Park regulates mitochondrial dynamics by inducing the degradation of the mitochondrial fusion protein Mitofusin/Marf in lrpprc2 mutants. In our genetic screen, we also discover that Bendless, a K63-linked E2 conjugase, is a regulator of Marf, as loss of bendless results in increased Marf levels. We show that Bendless is required for PINK1 stability, and subsequently for PINK1-Park mediated Marf degradation under physiological conditions, and in response to mitochondrial stress as seen in lrpprc2. Additionally, we show that loss of bendless in lrpprc2 mutant eyes results in photoreceptor degeneration, indicating a neuroprotective role for Bendless-PINK1-Park mediated Marf degradation. Based on our observations, we propose that certain forms of mitochondrial stress activate Bendless-PINK1-Park to limit mitochondrial fusion, which is a cell-protective response.

Buffered EGFR signaling regulated by spitz-to-argos expression ratio is a critical factor for patterning the Drosophila eye.

The Epidermal Growth Factor Receptor (EGFR) signaling pathway plays a critical role in regulating tissue patterning. Drosophila EGFR signaling achieves specificity through multiple ligands and feedback loops to finetune signaling outcomes spatiotemporally. The principal Drosophila EGF ligand, cleaved Spitz, and the negative feedback regulator, Argos are diffusible and can act both in a cell autonomous and non-autonomous manner. The expression dose of Spitz and Argos early in photoreceptor cell fate determination has been shown to be critical in patterning the Drosophila eye, but the exact identity of the cells expressing these genes in the larval eye disc has been elusive. Using single molecule RNA Fluorescence in situ Hybridization (smFISH), we reveal an intriguing differential expression of spitz and argos mRNA in the Drosophila third instar eye imaginal disc indicative of directional non-autonomous EGFR signaling. By genetically tuning EGFR signaling, we show that rather than absolute levels of expression, the ratio of expression of spitz-to-argos to be a critical determinant of the final adult eye phenotype. Proximate effects on EGFR signaling in terms of cell cycle and differentiation markers are affected differently in the different perturbations. Proper ommatidial patterning is robust to thresholds around a tightly maintained wildtype spitz-to-argos ratio, and breaks down beyond. This provides a powerful instance of developmental buffering against gene expression fluctuations.

Is basal cisternostomy in traumatic brain injury a need of hour or white elephant - A randomized trial to answer.

Background: Basal cisternostomy (BC) recently emerged as an adjuvant/alternative procedure to decompressive craniectomy (DC) in traumatic brain injuries (TBIs) with its potential to effectively reduce both intracranial pressure (ICP) and brain edema. However, its role in TBI is not yet established in the true sense and with clarity. The objective of the present study was to evaluate the effect of adjuvant BC on ICP, mortality, and clinicoradiological outcome. Methods: A single-center randomized control trial was conducted. Fifty patients were assigned to each DC-group and DC+BC-group. Randomization was done using the sealed envelope method. Both groups were followed in the postoperative period to compare the impact of surgery on ICP, radiological changes, and clinical outcome (mortality, days on ventilator/in intensive care unit (ICU), and Glasgow outcome scale-extended (GOS-E) at 12 weeks). Results: Both groups were comparable in terms of preoperative clinicoradiological characteristics. On postoperative days 1, 2, and 3, mean ICP was significantly low in the DC+BC-group (P < 0.0001). The decline in ICP in the DC+BC-group was significant in both moderate and severe TBI patients. In comparison, DC+BC-group has a shorter duration of mechanical ventilation/ICU stay and significantly better GOS-E score at 12 weeks (P < 0.0001*). The mortality rate was less in the DC+BC-group (48%) as compared to the DC-group (64%). Among radiological features, mean midline shift and mean outward brain herniation were significantly less in the DC+BC group. Bone-flap replacement was possible in ten patients of DC+BC-group at the time of primary surgery. Conclusion: Results of our study indicated that BC is beneficial in reducing both ICP and brain edema, which translates into favorable clinicoradiological outcomes.

Comparative Outcome Analysis of Endoscopic Third Ventriculostomy and Ventriculoperitoneal Shunt Surgery in Pediatric Hydrocephalus: An Experience of a Tertiary Care Center.

Background Endoscopic third ventriculostomy (ETV) and ventriculoperitoneal shunt surgery (VPS) are used for the surgical management of pediatric hydrocephalus. There is controversy regarding the safety and efficacy of these procedures according to age, etiologies, and type of hydrocephalus. Objective The purpose of this study was to compare the outcomes and complications of ETV and VPS in pediatric hydrocephalus and to evaluate the better procedure. Material and Methods We retrospectively analyzed the pediatric hydrocephalus cases that were operated by ETV and VPS at our department from June 2016 to June 2019. Data were analyzed with respect to the etiology of hydrocephalus, age, and gender of the patients. We compared the outcomes (success and failure) depending on age at surgery, etiology, and type of hydrocephalus, complications at 12 months of follow-up. Fisher's exact test and chi-square test were applied to test the significance of difference. Results There were 195 pediatric hydrocephalus cases, which were operated by ETV ( n = 43; 22.05%) and VPS ( n = 152; 77.95%). The mean age of the cases was 53.63 ± 60.24 (ranged 0.5-204 months) in the ETV group and 53.44 ± 54.10 (ranged 0.3-210 months) in the VPS group. The male-to-female ratio was 1.41:1 in the ETV group and 1.21: 1 in the VPS group. Overall, ETV had successful outcomes in 30(69.77%) cases and VPS in 102(67.11%) cases at 12 months of follow-up. The complication rates were found in 7 (16.28%) cases in the ETV group and 38 (25.0%) cases in the VPS group. At initial 0.5 months of follow-up, ETV required revisions in 6 (13.95%) cases and VPS in 15 (9.87%) cases. Conclusion Although the success and failure of both the procedure vary according to the age at surgery, etiology and type of hydrocephalus, and complications, and failure rates on follow-up, there were no statistically significant differences between the two surgical groups in respect to these variables.

Nanoengineered Ink for Designing 3D Printable Flexible Bioelectronics.

Flexible electronics require elastomeric and conductive biointerfaces with native tissue-like mechanical properties. The conventional approaches to engineer such a biointerface often utilize conductive nanomaterials in combination with polymeric hydrogels that are cross-linked using toxic photoinitiators. Moreover, these systems frequently demonstrate poor biocompatibility and face trade-offs between conductivity and mechanical stiffness under physiological conditions. To address these challenges, we developed a class of shear-thinning hydrogels as biomaterial inks for 3D printing flexible bioelectronics. These hydrogels are engineered through a facile vacancy-driven gelation of MoS2 nanoassemblies with naturally derived polymer-thiolated gelatin. Due to shear-thinning properties, these nanoengineered hydrogels can be printed into complex shapes that can respond to mechanical deformation. The chemically cross-linked nanoengineered hydrogels demonstrate a 20-fold rise in compressive moduli and can withstand up to 80% strain without permanent deformation, meeting human anatomical flexibility. The nanoengineered network exhibits high conductivity, compressive modulus, pseudocapacitance, and biocompatibility. The 3D-printed cross-linked structure demonstrates excellent strain sensitivity and can be used as wearable electronics to detect various motion dynamics. Overall, the results suggest that these nanoengineered hydrogels offer improved mechanical, electronic, and biological characteristics for various emerging biomedical applications including 3D-printed flexible biosensors, actuators, optoelectronics, and therapeutic delivery devices.

Keyhole approach in anterior circulation aneurysm: Current indication, advantages, technical limitations, complications and their avoidance.

Keyhole surgery recently evolved as a minimal invasive surgical approach for treatment of anterior circulation aneurysm. This review was done to evaluate the keyhole approach for anterior circulation aneurysms, their indications, advantages, technical limitations, complications and their avoidance. The literature review was performed with the phrase "keyhole approach for anterior circulation aneurysm" as a search term in PubMed central, Medline, Google scholar and Embase data base to identify all the articles published till December 2020. Out of 113 articles searched, 22 were included in this review after screening for eligibility. On analyzing these articles, there was total 2058 aneurysm in 1871 patients. Out of 2058 aneurysm, 988 were ruptured and 547 unruptured. In 5 studies, which include 344 aneurysms in 344 cases, aneurysm ruptured or unruptured status was not specified. The most frequent aneurysm site was anterior communicating artery (n=573). The size of the aneurysm mentioned in most of the study was <15 mm. The rate of complete occlusion was ranged from 93.6-100%. The range of intra operative rupture (IOR) was 0-28.6%. The mean operative time was ranged from 70 min-5.34 hours as reported in 13 studies. Good outcome [Glasgow outcome scale (GOS): 4-5] were seen in 75-100% cases. The frontalis muscle weakness has been reported in 3 studies and ranged from 0-1.99%. Keyhole surgery can be a safe and effective treatment modality for treatment of a selected anterior circulation aneurysm. In the experienced hand it has certain advantages over standard pterional craniotomy.

2D Covalent Organic Framework Direct Osteogenic Differentiation of Stem Cells.

2D covalent organic frameworks (COFs) are an emerging class of crystalline porous organic polymers with a wide-range of potential applications. However, poor processability, aqueous instability, and low water dispersibility greatly limit their practical biomedical implementation. Herein, a new class of hydrolytically stable 2D COFs for sustained delivery of drugs to direct stem cell fate is reported. Specifically, a boronate-based COF (COF-5) is stabilized using amphiphilic polymer Pluronic F127 (PLU) to produce COF-PLU nanoparticles with thickness of ≈25 nm and diameter ≈200 nm. These nanoparticles are internalized via clathrin-mediated endocytosis and have high cytocompatibility (half-inhibitory concentration ≈1 mg mL-1 ). Interestingly, the 2D COFs induce osteogenic differentiation in human mesenchymal stem cells, which is unique. In addition, an osteogenic agent-dexamethasone-is able to be loaded within the porous structure of COFs for sustained delivery which further enhances the osteoinductive ability. These results demonstrate for the first time the fabrication of hydrolytically stable 2D COFs for sustained delivery of dexamethasone and demonstrate its osteoinductive characteristics.

Enantioselective vinylogous aldol reaction of acylphosphonates with 3-alkylidene oxindoles.

A simple strategy for yielding chiral tertiary α-hydroxy phosphonates that integrates two highly biologically relevant scaffolds namely 3-alkylidene-2-oxindoles and phosphonates has been described. The hydrogen bonding ability of the bifunctional thiourea catalyst allows simultaneous dual activation of a vinylogous oxindole nucleophile and an acylphosphonate electrophile, affording hydroxyphosphonato-3-alkylidene-2-oxindoles as aldol adducts in high yields (up to 92%) with excellent stereocontrol (up to 99% ee).

Molecular cancer cell responses to solid compressive stress and interstitial fluid pressure.

Alterations to the mechanical properties of the microenvironment are a hallmark of cancer. Elevated mechanical stresses exist in many solid tumors and elicit responses from cancer cells. Uncontrolled growth in confined environments gives rise to elevated solid compressive stress on cancer cells. Recruitment of leaky blood vessels and an absence of functioning lymphatic vessels causes a rise in the interstitial fluid pressure. Here we review the role of the cancer cell cytoskeleton and the nucleus in mediating both the initial and adaptive cancer cell response to these two types of mechanical stresses. We review how these mechanical stresses alter cancer cell functions such as proliferation, apoptosis, and migration.

Light-Triggered In Situ Gelation of Hydrogels using 2D Molybdenum Disulfide (MoS2 ) Nanoassemblies as Crosslink Epicenter.

Light-responsive biomaterials are an emerging class of materials used for developing noninvasive, noncontact, precise, and controllable biomedical devices. Long-wavelength near-infrared (NIR) radiation is an attractive light source for in situ gelation due to its higher penetration depth and minimum side effects. The conventional approach to obtain crosslinked biomaterials relies heavily on the use of a photoinitiator by generating reactive species when exposed to short-wavelength radiation, which is detrimental to surrounding cells and tissue. Here, a new class of NIR-triggered in situ gelation system based on defect-rich 2D molybdenum disulfide (MoS2 ) nanoassemblies and thiol-functionalized thermoresponsive polymer in the absence of a photoinitiator is introduced. Exposure to NIR radiation activates the dynamic polymer-nanomaterials interactions by leveraging the photothermal characteristics of MoS2 and intrinsic phase transition ability of the thermoresponsive polymer. Specifically, upon NIR exposure, MoS2 acts as a crosslink epicenter by connecting with multiple polymeric chains via defect-driven click chemistry. As a proof-of-concept, the utility of NIR-triggered in situ gelation is demonstrated in vitro and in vivo. Additionally, the crosslinked gel exhibits the potential for NIR light-responsive release of encapsulated therapeutics. These light-responsive biomaterials have strong potential for a range of biomedical applications, including artificial muscle, smart actuators, 3D/4D printing, regenerative medicine, and therapeutic delivery.

Mitochondrial calcium at the synapse.

Mitochondria are dynamic organelles, which serve various purposes, including but not limited to the production of ATP and various metabolites, buffering ions, acting as a signaling hub, etc. In recent years, mitochondria are being seen as the central regulators of cellular growth, development, and death. Since neurons are highly specialized cells with a heavy metabolic demand, it is not surprising that neurons are one of the most mitochondria-rich cells in an animal. At synapses, mitochondrial function and dynamics is tightly regulated by synaptic calcium. Calcium influx during synaptic activity causes increased mitochondrial calcium influx leading to an increased ATP production as well as buffering of synaptic calcium. While increased ATP production is required during synaptic transmission, calcium buffering by mitochondria is crucial to prevent faulty neurotransmission and excitotoxicity. Interestingly, mitochondrial calcium also regulates the mobility of mitochondria within synapses causing mitochondria to halt at the synapse during synaptic transmission. In this review, we summarize the various roles of mitochondrial calcium at the synapse.

Two-dimensional metal organic frameworks for biomedical applications.

Two-dimensional (2D) metal organic frameworks (MOFs), are an emerging class of layered nanomaterials with well-defined structure and modular composition. The unique pore structure, high flexibility, tunability, and ability to introduce desired functionality within the structural framework, have led to potential use of MOFs in biomedical applications. This article critically reviews the application of 2D MOFs for therapeutic delivery, tissue engineering, bioimaging, and biosensing. Further, discussion on the challenges and strategies in next generation of 2D MOFs are also included. This article is categorized under: Nanotechnology Approaches to Biology > Nanoscale Systems in Biology.

Stereoselective formal [3 + 3] annulation of 3-alkylidene-2-oxindoles with ß,γ-unsaturated α-keto esters.

1,4-Diazabicyclo[2.2.2]octane (DABCO)-catalyzed [3 + 3] cycloaddition reaction of 3-alkylidene-2-oxindole and ß,γ-unsaturated α-keto esters under mild reaction conditions afforded the spirocyclohexene-oxindole with excellent diastereoselectivity. The [3 + 3] annulation is found to proceed through a vinylogous Michael-aldol cascade reaction and it allows rapid access to a diverse set of highly functionalized spirocyclohexene-oxindoles. Also, a bioactivity study of the compounds on mammalian sarcoma cells has reflected cell growth inhibitory/anti-cancer properties.

Photothermal modulation of human stem cells using light-responsive 2D nanomaterials.

Two-dimensional (2D) molybdenum disulfide (MoS2) nanomaterials are an emerging class of biomaterials that are photoresponsive at near-infrared wavelengths (NIR). Here, we demonstrate the ability of 2D MoS2 to modulate cellular functions of human stem cells through photothermal mechanisms. The interaction of MoS2 and NIR stimulation of MoS2 with human stem cells is investigated using whole-transcriptome sequencing (RNA-seq). Global gene expression profile of stem cells reveals significant influence of MoS2 and NIR stimulation of MoS2 on integrins, cellular migration, and wound healing. The combination of MoS2 and NIR light may provide new approaches to regulate and direct these cellular functions for the purposes of regenerative medicine as well as cancer therapy.

Management of multiloculated hydrocephalus in children with emphasis on role of CT ventriculography.

OBJECTIVE: The authors performed a retrospective review of children diagnosed with multiloculated hydrocephalus (MLH) in our institute. The goal was to analyze the different diagnostic and therapeutic modalities used with special emphasis on CT ventriculography (CTV). METHODS: Male and female patients below the age of 18 years diagnosed with MLH were included. Cases of uniloculated hydrocephalus like entrapped temporal horn or isolated fourth ventricle were excluded. We used iohexol for CTV and gadodiamide for MR ventriculography. Neuroendoscopic procedures performed were endoscopic fenestration, endoscopic third ventriculostomy (ETV), endoscopic septostomy, endoscopic aqueductoplasty, or a combination of the above. The cohort was divided into two groups (endoscopic or shunt) based on initial surgical intervention. RESULTS: A total of 52 patients were included, with 43 boys and 9 girls. The average age of presentation was 7.7 months. The most common predisposing factor for MLH was neonatal meningitis seen in 30 patients. Mean duration of follow-up was 39 months. CTV was used in 26 patients and MR ventriculography in three patients. In one patient, the diagnosis of MLH was ruled out after ventriculography. Patients who underwent ETV only had the best outcome with 71.4% success rate. At the end of follow-up, 14 patients (27%) were shunt independent. CONCLUSIONS: The present study indicates that CTV helps to accurately define the anatomy of the ventricles and determine the site of physiological CSF obstruction. This helps in therapeutic planning and in avoiding misdiagnoses. Further, neuroendoscopy has the potential to lead to shunt independence in some patients.

Nanoengineered Osteoinductive Bioink for 3D Bioprinting Bone Tissue.

Bioprinting is an emerging additive manufacturing approach to the fabrication of patient-specific, implantable three-dimensional (3D) constructs for regenerative medicine. However, developing cell-compatible bioinks with high printability, structural stability, biodegradability, and bioactive characteristics is still a primary challenge for translating 3D bioprinting technology to preclinical and clinal models. To overcome this challenge, we developed a nanoengineered ionic covalent entanglement (NICE) bioink formulation for 3D bone bioprinting. The NICE bioinks allow precise control over printability, mechanical properties, and degradation characteristics, enabling custom 3D fabrication of mechanically resilient, cellularized structures. We demonstrate cell-induced remodeling of 3D bioprinted scaffolds over 60 days, demonstrating deposition of nascent extracellular matrix proteins. Interestingly, the bioprinted constructs induce endochondral differentiation of encapsulated human mesenchymal stem cells (hMSCs) in the absence of osteoinducing agent. Using next-generation transcriptome sequencing (RNA-seq) technology, we establish the role of nanosilicates, a bioactive component of NICE bioink, to stimulate endochondral differentiation at the transcriptome level. Overall, the osteoinductive bioink has the ability to induce formation of osteo-related mineralized extracellular matrix by encapsulated hMSCs in growth factor-free conditions. Furthermore, we demonstrate the ability of NICE bioink to fabricate patient-specific, implantable 3D scaffolds for repair of craniomaxillofacial bone defects. We envision development of this NICE bioink technology toward a realistic clinical process for 3D bioprinting patient-specific bone tissue for regenerative medicine.

Endoscopic Transcortical Transventricular Management of Cystic Craniopharyngioma: Outcome Analysis of 32 Cases at a Tertiary Care Center.

BACKGROUND: Microsurgical resection has been considered the gold standard treatment of craniopharyngioma, but lately, it has found less favor due to its morbidity and is being replaced by minimally invasive cyst drainage procedures. We present our experience of transventricular endoscopy and cyst drainage along with its technique and have analyzed its results. MATERIALS AND METHODS: Clinical and radiological data of all cystic craniopharyngioma patients treated by transventricular endoscopic cyst drainage and Ommaya placement were retrieved and analyzed. RESULTS: Thirty-two patients underwent endoscopic cyst drainage during the study period. All patients had immediate clinical and radiological improvement. No significant complications were seen. All patients underwent adjuvant radiotherapy and six patients (18.7%) showed recurrence. Three patients died in the follow-up period. CONCLUSIONS: Endoscopic transcortical transventricular cyst drainage with Ommaya reservoir along with adjuvant radiotherapy is a simple, safe, and effective treatment modality.