New Tmc1 Deafness Mutations Impact Mechanotransduction in Auditory Hair Cells

Transmembrane channel-like protein isoform 1 (TMC1) is a major component of the mechano-electrical transducer (MET) channel in cochlear hair cells and is subject to numerous mutations causing deafness. We report a new dominant human deafness mutation, TMC1 p.T422K, and have characterized the homologous mouse mutant, Tmc1 p.T416K, which caused deafness and outer hair cell (OHC) loss by the fourth postnatal week. MET channels showed decreased Ca²⁺ permeability and resting open probability, but no change in single-channel conductance or expression. Three adjacent deafness mutations are TMC1 p.L416R, p.G417R, and p.M418K, the last homologous to the mouse Beethoven that exhibits similar channel effects. All substitute a positive for a neutral residue, which could produce charge screening in the channel pore or influence binding of an accessory subunit. Channel properties were compared in mice of both sexes between dominant (Tmc1 p.T416K, Tmc1 p.D569N) and recessive (Tmc1 p.W554L, Tmc1 p.D528N) mutations of residues near the putative pore of the channel. Tmc1 p.W554L and p.D569N exhibit reduced maximum current with no effect on single-channel conductance, implying a smaller number of channels transported to the stereociliary tips; this may stem from impaired TMC1 binding to LHFPL5. Tmc1 p.D528N, located in the pore''s narrowest region, uniquely caused large reductions in MET channel conductance and block by dihydrostreptomycin (DHS). For Tmc1 p.T416K and Tmc1 p.D528N, transduction loss occurred between P15 and P20. We propose two mechanisms linking channel mutations and deafness: decreased Ca²⁺ permeability, common to all mutants, and decreased resting open probability in low Ca²⁺, confined to dominant mutations.SIGNIFICANCE STATEMENT Transmembrane channel-like protein isoform 1 (TMC1) is thought to be a major component of the mechanotransducer channel in auditory hair cells, but the protein organization and channel structure are still uncertain. We made four mouse lines harboring Tmc1 point mutations that alter channel properties, causing hair cell degeneration and deafness. These include a mouse homolog of a new human deafness mutation pT416K that decreased channel Ca²⁺ permeability by introducing a positively-charged amino acid in the putative pore. All mutations are consistent with the channel structure predicted from modeling, but only one, p.D528N near the external face of the pore, substantially reduced channel conductance and Ca²⁺ permeability and virtually abolished block by dihydrostreptomycin (DHS), strongly endorsing its siting within the pore.     

Chest radiography requirements for patients with asymptomatic COVID-19 undergoing coronary artery bypass surgery: Three case reports

BACKGROUNDThe coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus-2, represents a major challenge to health care systems both globally and regionally, with many opting by cancelling elective surgeries. Cardiac operations in patients diagnosed with COVID-19 have been imperative due to their emergency nature, critical condition of patients awaiting cardiac surgery, and accumulated number of cardiac surgical interventions throughout the last months.CASE SUMMARYHere we describe three COVID-19 positive cases who underwent coronary surgery, on an urgent basis. We did not experience worsening of the patients’ clinical condition due to COVID-19 and therefore a routine post-operative chest X-ray (CXR) was not required. None of the health care providers attending the patients endured cross infection. Further trials would be needed in order to confirm these results.CONCLUSIONWhile the pandemic has adversely hit the health systems worldwide, cardiac surgical patients who concomitantly contracted COVID-19 may undergo a smooth post-operative course as a routine post-operative CXR may not be required.     

Fracture resistance of simulated immature roots using Biodentine and fiber post compared with different canal-filling materials under aging conditions

Objective

The aim of this study was to evaluate the fracture resistance of simulated immature roots using Biodentine (BD) and fiber post (FP) compared with different root canal-filling materials under aging conditions.

Materials and methods

One hundred and forty maxillary central anterior teeth were randomly divided into seven groups (n = 20/group). Negative control received no treatment. In the other groups, the root canals were prepared to simulate immature teeth. The root canals were filled with a 4-mm apical plug of BD and restored intraradicular with BD, BD + FP, composite resin (CR), CR + FP, and gutta-percha (GP). Positive controls were instrumented but without restoration. Teeth were subjected to thermocycling and received cyclic loading before fracture resistance test. Fracture resistance was conducted using a universal testing machine with a crosshead speed of 0.5 mm/min until fracture. Load to fracture was recorded in newtons (N). Data were statistically analyzed using one-way analysis of variance and Tukey’s test at P < 0.05.

Results

Root canals restored intraradicular with BD + FP and CR + FP showed the highest fracture resistance compared with the other experimental groups (P < 0.001). There was no significant difference in the fracture resistance between CR and BD groups (P = 0.998). GP and positive control groups were significantly lower resistance to fracture than the other groups (P < 0.001).

Conclusions

Intraradicular reinforcement with BD + FP and CR + FP enhanced the fracture resistance of simulated immature teeth than the other experimental groups.

Clinical relevance

Biodentine or composite resin combined with fiber post could be used to reinforce immature teeth with an apical Biodentine plug.

     

Ilimaquinone (marine sponge metabolite) as a novel inhibitor of SARS-CoV-2 key target proteins in comparison with suggested COVID-19 drugs: designing,docking and molecular dynamics simulation study

The outbreak of novel coronavirus, SARS-CoV-2, has infected more than 36 million people and caused approximately 1 million deaths around the globe as of 9 October 2020. The escalating outspread of the virus and rapid rise in the number of cases require the instantaneous development of effectual drugs and vaccines. Presently, there are no approved drugs or vaccine available to treat the infection. In such scenario, one of the propitious therapeutic approaches against viral infection is to explore enzyme inhibitors amidst natural compounds, utilizing computational approaches aiming to get products with negligible side effects. In the present study, the inhibitory prospects of ilimaquinone (marine sponge metabolite) were assessed in comparison with hydroxychloroquine, azithromycin, favipiravir, ivermectin and remdesivir at the active binding pockets of nine different vital SARS-CoV-2 target proteins (spike receptor binding domain, RNA-dependent RNA polymerase, Nsp10, Nsp13, Nsp14, Nsp15, Nsp16, main protease, and papain-like-protease), employing an in silico molecular interaction based approach. In addition, molecular dynamics (MD) simulations of the SARS-CoV-2 papain-like protease (PLpro)–ilimaquinone complex were also carried out to calculate various structural parameters including root mean square fluctuation (RMSF), root mean square deviation (RMSD), radius of gyration (R_g) and hydrogen bond interactions. PLpro is a promising drug target, due to its imperative role in viral replication and additional function of stripping ubiquitin and interferon-stimulated gene 15 (ISG15) from host-cell proteins. In light of the possible inhibition of all vital SARS-CoV-2 target proteins, our study has emphasized the importance to study in depth ilimaquinone actions in vivo.

Inhibitory potential of ilimaquinone (marine sponge metabolite) against nine essential SARS-CoV-2 target proteins, employing a molecular interaction and dynamics simulation approach.     

Design and formulation of polymeric nanosponge tablets with enhanced solubility for combination therapy

Three drugs namely caffeine, paracetamol, and aceclofenac are commonly used for treating various acute and chronic pain related ailments. These 3 drugs have varied solubility profiles, and formulating them into a single tablet did not have the desired dissolution profile for drug absorption. The objective of the present research was to tailor the drug release profile by altering drug solubility. This was achieved by loading the drug into nanosponges. Here, three-dimensional colloidal nanosponges were prepared using β-cyclodextrin with dimethyl carbonate as a cross-linker using the hot-melt compression method. The prepared nanosponges were characterized by FTIR, ¹H NMR spectroscopy, DSC, XRPD studies and SEM. The FTIR and DSC results obtained indicated polymer-drug compatibility. The ¹H NMR spectroscopy results obtained indicated the drug entrapment within nanosponges with the formation of the inclusion complex. XRPD studies showed that the loaded drug had changed crystalline properties altering drug solubility. SEM photographs revealed the porous and spongy texture on the surface of the nanosponge. Box–Behnken experimental design was adopted for the optimization of nanosponge synthesis. Among the synthesized nanosponges containing paracetamol, aceclofenac and caffeine, batch F3–P31, F3–A31 and F3–C31 were considered optimized. Their particle size was 185, 181 and 199 nm with an entrapment efficiency of 81.53, 84.96, and 89.28% respectively. These optimized nanosponges were directly compressed into tablets and were studied for both pre and post-compression properties including in vitro drug release. The prepared tablet showed desired drug dissolution properties compared to the pure drug. The above outcomes indicated the applicability of nanosponges in modulating the drug release with varied solubility for combination therapy.

Polymeric nanosponges as potential carriers for successful combination therapy of poorly soluble drugs (paracetamol, aceclofenac, caffeine).     

Identification of potential inhibitors of Zika virus NS5 RNA-dependent RNA polymerase through virtual screening and molecular dynamic simulations

Zika virus (ZIKV) is one of the mosquito borne flavivirus with several outbreaks in past few years in tropical and subtropical regions. The non-structural proteins of flaviviruses are suitable active targets for inhibitory drugs due to their role in pathogenicity. In ZIKV, the non-structural protein 5 (NS5) RNA-Dependent RNA polymerase replicates its genome. Here we have performed virtual screening to identify suitable ligands that can potentially halt the ZIKV NS5 RNA dependent RNA polymerase (RdRp). During this process, we searched and screened a library of ligands against ZIKV NS5 RdRp. The selected ligands with significant binding energy and ligand-receptor interactions were further processed. Among the selected docked conformations, top five was further optimized at atomic level using molecular dynamic simulations followed by binding free energy calculations. The interactions of ligands with the target structure of ZIKV RdRp revealed that they form strong bonds within the active sites of the receptor molecule. The efficacy of these drugs against ZIKV can be further analyzed through in-vitro and in-vivo studies.