Novel Management of Hypersensitive Dentin Using Propolis-based Herbal Desensitizing Agents: An In Vitro Scanning Electron Microscopic Study.

AIM: The objective of this in vitro study was to assess the efficacy of novel propolis-based varnish against the two conventional varnishes on quantitative and qualitative assessments of occlusion of dentin tubules and resistance to erosive and abrasive wears employing scanning electron microscope (SEM). METHODS: Thirty human premolars free from caries extracted due to orthodontic reasons were included in the study. Experimental group was done based on treatment received and divided into three groups. Group A: ClinProXT Varnish (n = 10), Group B: MI Varnish (n = 10), and Group C: Propolis Varnish (n = 10) were applied. Teeth were cleaned and decoronation of crown was done with dentin disks. Dentin specimens of dimension 4 × 4 × 2 mm were prepared and subjected to finishing and polishing. The sample specimens were submersed in EDTA solution for a period of five minutes to open up the dentinal tubules. This was followed by treatment with varnishes and subjection to acidic-abrasive challenge. The specimens were analyzed with an image analyzer connected to SEM for the verification of the number of opened dentin tubules. The parameter assessed in SEM includes size, topography and surface characteristics of dentinal tubule were assessed. The obliteration potential of dentinal tubules was assessed with SEM images. Additionally, the dentin surface loss and resistance to acidic and abrasive wear were also evaluated with SEM. Data were analyzed with two-way analysis of variance (ANOVA) with post hoc Tukey's test. RESULTS: MI Varnish caused higher obliteration of dentin tubules followed by ClinproXT Varnish. Propolis Varnish showed the least obliteration of dentinal tubules among tested experimental groups. After acidic-abrasive challenge, Propolis Varnish was found to be more efficient with less material loss among the experimental groups tested. There was an insignificant difference among the MI Varnish and ClinProXT Varnish groups. CONCLUSION: Simulation of hypersensitive lesions mimicking the clinical scenario was a challenging task in this in vitro study. All varnishes tested in the study had good efficacy in the management of dentin hypersensitivity (DH). Propolis-based varnish had good resistance to material loss after subjection to acidic-abrasive challenge among the tested materials. The casein phosphopeptide (CPP)-amorphous calcium phosphate (ACP)-based MI Varnish had good efficacy to obliterate the dentinal tubules among the tested materials. It was prudent to select the varnishes with good long-term efficacy to survive in the clinical scenario which still remains a challenging task for the clinicians. CLINICAL SIGNIFICANCE: The stability of the varnish plays a vital role in maintenance of its long-term efficacy. The chemical nature along with the ability of the material to interact with the substrate plays a major role in management of DH.

Clicking a Fish: Click Chemistry of Different Biomolecules in Danio rerio.

Click chemistry has found remarkable applications in imaging biomacromolecules in cellular systems and even organisms. Over the past few years, bioorthogonal click reactions have been improved and tailored for specific applications, allowing the selective labeling of biomolecules like DNA, RNA, proteins, glycans, and lipids. The zebrafish, Danio rerio, is a prime model organism for vertebrate development and has gained importance in experimental biology and biochemistry for its ease of handling and appreciable genetic homology to mammals. With respect to biomolecular labeling, the zebrafish represents the next level of complexity relative to in vitro cell line models, challenging researchers to further improve the specificity and targeting of their labeling approaches, and to reduce the level of interference with the host. At the same time, bioorthogonal labeling has facilitated an improved understanding of embryonic zebrafish development. Highlighted in this Perspective are click chemistry approaches that rely on (i) designing analogues of endogenous target molecules, (ii) utilizing the conjugation moiety on these analogues to attach fluorophores by clicking them outside or inside living systems, and finally, (iii) imaging the cells or tissue or, sometimes, the whole organism. While glycans and proteins have been extensively studied in D. rerio using bioorthogonal click chemistry for their intimate involvement in embryonic development, DNA and RNA remain less investigated. Together with the formulation of strategies to perform click reactions with the weakest impact on the host system, bioorthogonal approaches have enormous potential to visualize organisms in all their molecular splendor.

microRNA-145 downregulates SIP1-expression but differentially regulates proliferation, migration, invasion and Wnt signaling in SW480 and SW620 cells.

microRNA-145 (miR-145) has been shown to act as a tumor suppressor in colorectal cancer but its role in the regulation of epithelial-mesenchymal transition (EMT) is unclear. Ectopic expression of miR-145 suppressed the proliferation, migration and invasion in SW480 but surprisingly enhanced these traits in its metastatic counterpart, SW620 cells, while, antimiR-145 reversed the effects of miR-145 in both of these human colorectal cancer cells. In SW480 and SW620 cells, SMAD-interacting protein 1 (SIP1), was identified as a target of miR-145, and its expression was suppressed both at mRNA and protein levels, and siRNA-SIP1 mimicked the effects of miR-145. Further, re-introduction of SIP1 alone or its co-expression with miR-145, rescued SW480 and SW620 cells from the effects of miR-145, indicating that the distinct functions of miR-145 might be mediated, in part, through SIP1. Since Wnt signaling plays an essential role in EMT in CRC progression, the effects of miR-145 on the expression of Wnt signaling intermediates and EMT markers were studied. Re-expression of miR-145 was found to downregulate the expression of CTNNB1, TCF4, CCND1, VIM, and SNAI, but, upregulated CDH1 expression in SW480 cells. On the other hand, miR-145 exhibited an oncogenic potential in SW620 cells by actuating Wnt signaling and the expression of EMT-relevant markers. These results strongly hint that the paradoxical functions of miR-145 in the regulation of proliferation, migration and invasion might be mediated through downregulation of SIP1, and differential tuning of Wnt signaling and EMT-mediators.

miR-214 activates TP53 but suppresses the expression of RELA, CTNNB1, and STAT3 in human cervical and colorectal cancer cells.

High Mobility Group AT-hook 1 (HMGA1) was identified as a target of miR-214 in human cervical and colorectal cancers (CaCx and CRC) in a previous study. While the expression of miR-214 remains suppressed, HMGA1 behaves as a potent oncogene and plays crucial roles in several aberrant signalling pathways by interacting with intermediates like RELA, CTNNB1, STAT3, and TP53 in CaCx and CRC. Hypothetically, miR-214 should be able to regulate the stabilization of some of these intermediates through the regulation of HMGA1. This was assessed by ectopically expressing miR-214 or complementarily, by inhibiting the expression of HMGA1. In promoter luciferase assays, miR-214 inhibited NF-κB and Wnt activities but elevated TP53 activity in cancer cells. Further, miR-214 suppressed the expression of HMGA1, RELA, CTNNB1, and STAT3 while elevating TP53 levels, similar to when small interfering RNA (siRNA) against HMGA1 was used, as revealed by Western blotting. It is suggested that poor expression of miR-214, commonly reported in CaCx and CRC tissues, may not only result in the sustained expression of HMGA1 but also that of RELA, CTNNB1, and STAT3, and a congruent suppression of TP53 during cancer initiation/progression. These several states are, however, reversed when miR-214 is reintroduced and could explain the tumour suppressive functions observed in earlier studies. Further studies are, however, required to reveal how microRNA-mediated regulation of HMGA1 expression may affect individual signalling pathways in CaCx and CRC. Current results reveal that miR-214 is not only able to regulate the expression of its direct target, HMGA1, but also that of a few signalling intermediates like TP53, RELA, CTNNB1, and STAT3, with which HMGA1 interacts. These intermediates play crucial roles in signalling pathways commonly deregulated in human CaCx and CRC. Hence, it is proposed that miR-214 might act as a tumour suppressor by regulating several aberrant signalling pathways through HMGA1. This knowledge has the potential to help design novel therapeutic strategies in CaCx and CRC.

MicroRNA-214 suppresses growth, migration and invasion through a novel target, high mobility group AT-hook 1, in human cervical and colorectal cancer cells.

BACKGROUND: MicroRNA-214 (miR-214) has been shown to act as a tumour suppressor in human cervical and colorectal cancer cells. The aim of this study was to experimentally validate high mobility group AT-hook 1 as a novel target for miR-214-mediated suppression of growth and motility. METHODS: HMGA1 and miR-214 expression levels were estimated in cervical and colorectal clinical specimens using qPCR. HMGA1 3' untranslated region luciferase assays were performed to validate HMGA1 as a target of miR-214. Effect of altering the expression of miR-214 or HMGA1 on proliferation, migration and invasion of human cervical and colorectal cancer cells was investigated. RESULTS: miR-214 expression was poor while that of HMGA1 was high in cervical and colorectal cancer tissues. miR-214-re-expression or HMGA1 downregulation inhibited proliferation, migration and invasion of cancer cells while miR-214 inhibition had opposite effects. miR-214 was demonstrated to bind to the wild-type 3' untranslated region of HMGA1 but not with its mutant. CONCLUSIONS: Low expression of miR-214 concurrent with elevated levels of HMGA1 may contribute to cervical and colorectal cancer progression. miR-214-mediated regulation of HMGA1 is a novel mechanism for its tumour-suppressive actions in human cervical and colorectal cancer cells and opens up avenues for novel therapeutic strategies for these two cancers.

microRNA-146a inhibits proliferation, migration and invasion of human cervical and colorectal cancer cells.

microRNAs (miRNAs) play significant roles in diverse biological processes and their deregulation is implicated in carcinogenesis. miR-146a executes tumour suppressive or oncogenic functions depending on the cancer type, but its effect on human cervical (CaCx) and colorectal (CRC) cancers have not been examined thus far. In this study, miR-146a exhibited high expression in CaCx but poor expression in CRC, in comparison to corresponding normal tissues. Nevertheless, ectopic expression of miR-146a inhibited proliferation in both CaCx and CRC cells and curbed their migration and invasion. When the expression of endogenous miR-146a was suppressed, proliferative, migratory and invasive capacities of CaCx and CRC cells increased, suggesting an anti-tumourigenic function for miR-146a. Re-expression of miR-146a down-regulated the expression of crucial signalling intermediates: CTNNB1, STAT3, RELA, CCND1 and SNAI1, and enhanced TP53 and CDH1 expression. Thus, the present study reveals a hitherto unknown tumour suppressive role for miR-146a providing a plausible mechanistic basis for it.

Downregulation of HMGB1 by miR-34a is sufficient to suppress proliferation, migration and invasion of human cervical and colorectal cancer cells.

High mobility group box 1 (HMGB1) is a ubiquitous nuclear protein known to be highly expressed in human cervical (CaCx) and colorectal (CRC) cancers, and sustained high levels of HMGB1 contribute to tumourigenesis and metastasis. HMGB1-targeted cancer therapy is of recent interest, and there are not many studies on miRNA-mediated HMGB1 regulation in these cancers. Since miRNA-based therapeutics for cancer is gaining importance in recent years, it was of interest to predict miRNAs targeting HMGB1. Based on the identification of a potential miR-34a response element in HMGB1-3' untranslated region (3'UTR) and an inverse correlation between HMGB1 and miR-34a expression levels in CaCx and CRC tissues, from a subset of the local population as well as a large sampling from TCGA database, experiments were performed to validate HMGB1 as a direct target of miR-34a in CaCx and CRC cells. Ectopic expression of miR-34a decreased the wild-type HMGB1-3'UTR luciferase activity but not that of its mutant in 3'UTR luciferase assays. While forced expression of miR-34a in CaCx and CRC cells inhibited HMGB1 mRNA and protein levels, proliferation, migration and invasion, inhibition of endogenous miR-34a enhanced these tumourigenic properties. siRNA-mediated HMGB1 suppression imitated miR-34a expression in reducing proliferation and metastasis-related events. Combined with the disparity in expression of miR-34a and HMGB1 in clinical specimens, the current findings would help in not only understanding the complexity of miRNA-target regulatory mechanisms but also in designing novel therapeutic interventions in CaCx and CRC.

Long-term follow up of single-stage anterior cruciate ligament reconstruction and high tibial osteotomy and its relation with posterior tibial slope.

BACKGROUND: Open-wedge high tibial osteotomy is considered to be an effective treatment for medial compartmental osteoarthritis. It is generally admitted that tibial slope increases after open-wedge high tibial osteotomy and decreases after closing-wedge high tibial osteotomy. Young patients with anterior cruciate ligament (ACL) deficiency along with medial compartment osteoarthritis need a combined procedure of ACL reconstruction along with high tibial osteotomy to regain physiological knee kinematics and to avoid chondral damage. MATERIALS AND METHODS: We retrospectively analysed data from 30 patients who underwent arthroscopic ACL reconstruction along with medial opening-wedge osteotomy from Jan 2004 to June 2012 with a minimum follow up of 2 years. The pre-operative and post-operative posterior tibial slopes were measured. Functional outcome was analysed using clinico-radiological criteria, IKDC scoring and Lysholm score. RESULTS: Post-operative patients improved both clinically and functionally. The patients who had posterior tibial slope >5° decrease, compared to patients who had less <5° decrease, had better functional scores (IKDC and Lysholm score), which was statistically significant (p < 0.05). CONCLUSION: Our study has shown that decreasing the tibial slope >5° compared to pre-operative value has functionally favourable effect on the reconstructed ACL graft and outcome. It is known that increasing slope causes an anterior shift in tibial resting position that is accentuated under axial loads. This suggests that decreasing tibial slope may be protective in an ACL deficient knee. Hence by placing the tricortical graft posterior to midline in the opening wedge reduces the posterior tibial slope and thereby reduces the stress on the graft leading to better functional outcome.

Design and realization of a novel haptic graspable interface for augmenting touch sensations.

A novel haptic grasper that renders touch sensations to the user in 3-DoF (degrees of freedom), namely linear, rotary, and grasping motions, is presented. The touch sensations of the grasper include the combination of kinesthetic and tactile modalities such as stiffness, texture, and shape. The device is equipped with two swappable modular segments that provide stiffness and shape sensations. To increase the haptic fidelity, the textural surfaces that surround the outer surface of the segments are equipped with vibro-actuators underneath them. These vibro-actuators contribute to increasing the number of perceivable textures by varying amplitude, frequency, duration, and envelope of vibrations. The proposed device is characterized in terms of stiffness, shape and texture rendering capabilities. The experimental results validate the effectiveness of the developed haptic grasper in virtual/remote interactions. Also, the user studies and statistical analysis demonstrate that the users could perceive the high-fidelity haptic feedback with the unified sensations of kinesthetic and tactile cues.

Racial Disparities in COVID-19 Hospitalization and In-hospital Mortality at the Height of the New York City Pandemic.

OBJECTIVES: Investigate whether or not race is associated with differences in hospitalization and survival to discharge among patients with coronavirus disease-2019 (COVID-19) at the height of the pandemic in New York City (NYC). METHODS: Single-center retrospective cohort study of COVID-19 patients hospitalized at our university-affiliated NYC hospital from 3/10/20 through 4/13/20 with follow-up to 5/1/20. Our primary endpoint was hospitalization rate among patients with confirmed COVID-19 compared with the regional population based on race. Our secondary endpoint survival to discharge among hospitalized COVID-19 patients. NYC Department of Health data were used to calculate hospitalization odds ratios. Chi-square and t tests were used to compare categorial and continuous variables, respectively. Cox proportional hazards regression and predictive analysis were used to investigate our endpoints further. RESULTS: Our cohort of 734 patients included 355 women (48.4%), 372 Blacks (50.7%), 214 Whites (29.2%), and 92 Hispanics (12.5%) in our analysis. Blacks were nearly twice as likely as Whites to require hospitalization for COVID-19 (OR 1.89, 95% CI, 1.59-2.24, p < 0.001). Hispanics were also more likely to suffer in-hospital mortality from COVID-19 compared with Whites (HR 1.84; 95% CI 1.21-2.80; p = 0.005). There was a non-significant increased hazard of in-hospital mortality among Blacks when compared with Whites (HR, 1.30; 95% CI, 0.95-1.78; p = 0.09). CONCLUSIONS AND RELEVANCE: Blacks were more likely than Whites to require hospitalization for COVID-19 while Hispanics were more likely to experience in-hospital mortality. Further investigation into the socioeconomic factors underlying racial disparities in COVID-19 survival and severity requiring hospitalization is needed on a national scale.

Design of a tether-driven minimally invasive robotic surgical tool with decoupled degree-of-freedom wrist.

BACKGROUND: Dexterous surgical tool wrists used in tele-operated robotic surgery typically have mechanically coupled pitch and yaw degree of freedom (DoF). This leads to complex control requirements. MATERIALS AND METHODS: The design of a robotic surgical tool with a mechanically decoupled dexterous wrist which uses stationary tether guides to guide drive tethers is presented. The tethers are routed through the plane of symmetry of the tool and follow law of belting to mechanically decouple the wrist. An optimization procedure for the placement of the stationary tether guides to minimize the change in tether length is presented. RESULTS: Experimental and analytical results confirm the decoupled motion capability of the wrist. Also, the change in length of tether segments over the operating range of motion was found to be very small. CONCLUSION: A prototype has been fabricated through metal 3D printing and integrated to a tele-operated robotic setup to demonstrate its utility in surgical application.

Bioorthogonal mRNA labeling at the poly(A) tail for imaging localization and dynamics in live zebrafish embryos.

Live imaging of mRNA in cells and organisms is important for understanding the dynamic aspects underlying its function. Ideally, labeling of mRNA should not alter its structure or function, nor affect the biological system. However, most methods applied in vivo make use of genetically encoded tags and reporters that significantly enhance the size of the mRNA of interest. Alternately, we utilize the 3' poly(A) tail as a non-coding repetitive hallmark to covalently label mRNAs via bioorthogonal chemistry with different fluorophores from a wide range of spectra without significantly changing the size. We demonstrate that the labeled mRNAs can be visualized in cells and zebrafish embryos, and that they are efficiently translated. Importantly, the labeled mRNAs acquired the proper subcellular localization in developing zebrafish embryos and their dynamics could be tracked in vivo.

microRNA-145 modulates epithelial-mesenchymal transition and suppresses proliferation, migration and invasion by targeting SIP1 in human cervical cancer cells.

PURPOSE: Previously, it has been reported that microRNA-145 (miR-145) is lowly expressed in human cervical cancers and that its putative tumour suppressive role may be attributed to epithelial-mesenchymal transition (EMT) regulation. Here, we aimed to assess whether miR-145 may affect EMT-associated markers/genes and suppress cervical cancer growth and motility, and to provide a mechanistic basis for these phenomena. METHODS: The identification of the SMAD-interacting protein 1 (SIP1) mRNA as putative miR-145 target was investigated using a 3' untranslated region (3'UTR) luciferase assay and Western blotting, respectively. The functional effects of exogenous miR-145 expression, miR-145 suppression or siRNA-mediated SIP1 expression down-regulation in cervical cancer-derived C33A and SiHa cells were analysed using Western blotting, BrdU incorporation (proliferation), transwell migration and invasion assays. In addition, the expression levels of miR-145 and SIP1 were determined in primary human cervical cancer and non-cancer tissue samples using qRT-PCR. RESULTS: We found that miR-145 binds to the wild-type 3'UTR of SIP1, but not to its mutant counterpart, and that, through this binding, miR-145 can effectively down-regulate SIP1 expression. In addition, we found that exogenous miR-145 expression or siRNA-mediated down-regulation of SIP1 expression attenuates the proliferation, migration and invasion of C33A and SiHa cells and alters the expression of the EMT-associated markers CDH1, VIM and SNAI1, whereas inhibition of endogenous miR-145 expression elicited the opposite effects. The expression of miR-145 in cervical cancer tissue samples was found to be low, while that of SIP1 was found to be high compared to non-cancerous cervical tissues. An inverse expression correlation between the two was substantiated through the anlaysis of data deposited in the TCGA database. CONCLUSION: Our data indicate that low miR-145 expression levels in conjunction with elevated SIP1 expression levels may contribute to cervical cancer development. MiR-145-mediated regulation of SIP1 provides a novel mechanistic basis for its tumour suppressive mode of action in human cervical cancer cells.

Re-expression of HPV16 E2 in SiHa (human cervical cancer) cells potentiates NF-κB activation induced by TNF-α concurrently increasing senescence and survival.

Re-expression of E2 in human papillomavirus (HPV) transformed tumour cells can induce apoptosis; however, some evidences also attribute an important role to E2 in sustaining tumorigenesis. In the present paper, we studied the effects of tumour necrosis factor (TNF)-α-mediated NF-κB (nuclear factor kappa-light-chain-enhancer of activated B-cells) activation on E2-induced senescence in HPV16-integrated SiHa cells. The results show that E2 inhibits endogenous E6 gene expression and sensitizes SiHa cells to TNF-α-induced NF-κB activation. Under this condition there was an increase in the expression of senescent proteins p53, p21, p27 and p16 and senescence-associated (SA)-ß-galactosidase activity indicating that TNF-α augments E2-mediated senescence. Re-expression of E2 expression with TNF-α treatment resulted in an increase in the expression of anti-apoptotic Bcl2 (B-cell lymphoma 2) protein and other pro-survival genes like cyclin D1 (cyc D1), survivin and hTERT (human telomerase reverse transcriptase). Concomitantly, E2 + TNF-α combination increased the survival of SiHa cells by positive changes in viability, proliferation and colony formation. E2-induced apoptotic tendency shifted towards senescence in presence of TNF-α by arresting the cells at both G0/G1 and G2/M phases, thus enhancing cell survival. Another observation in the present study is the significant up-regulation of key senescence messaging factors regulated by NF-κB namely interleukin (IL)-6, IL-8, high-mobility group protein A (HMGA)1 and B (HMGB)1 in E2-transfected cells treated with TNF-α. Our data provide a mechanistic basis and a new insight for the role of TNF-α and E2 in linking cellular senescence, tumorigenesis and HPV re-infection.