Karthik Subramanian Chandrasekaran

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.

microRNA-145 downregulates SIP1-expression but differentially regulates proliferation, migration, invasion and Wnt signalling 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.

Static balancing and inertia compensation of a master manipulator for tele-operated surgical robot application

Tele-operated robotic surgery is becoming popular and there is a need for design of better robotic arms for increased dexterity and user convenience. In the master-slave configuration of surgical robots, master manipulator plays an important role in ensuring user comfort and dexterity. The design and analysis of a master arm with 6 DOF with a wrist decoupled configuration is presented here. Primary focus of the design is to reduce the number of balancing masses required. By strategically positioning the joint axes, the balancing requirement for some of the axes has been eliminated. Also, through proper design modifications, remote placement of balancing masses has been achieved which reduces the net inertia of the system. Static balancing of the arm is analysed and an optimal design to achieve the balancing is presented. The residual imbalance is corrected through inertia compensation using feed forward control. The design details of the arm, its kinematic analysis, and balancing are presented in this paper.

MagNex — Expendable robotic surgical tooltip

This paper presents the design of a single use disposable compliant surgical tooltip for a tele-operated surgical robot. The proposed design aims at mitigating bio-fouling of surgical tools. By implementing a monolithic design for surgical tooltip, the cleaning and sterilization processes needed after every surgery for traditional surgical robotic tools is considerably simplified. The proposed design has 3 degrees of freedom (DOF) with a modified serpentine joint for enhanced buckling strength and off-axis stiffness. A magnetic force based coupling is proposed as a means of transferring power through a hermetic barrier to maneuver the tooltip. The tooltip is thus coupled through a hermetically sealed tool shaft preventing biological material from entering the tool shaft. By having a pluggable tooltip, modularity is also achieved by interchangeable tooltip instead of replacing the whole tool during surgery. A prototype of the proposed design is developed and its functional performance is validated. Owing to the magnetic nexus which forms the magnetic coupling, the tool is named MagNex.

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Reply: Comment on ‘ MicroRNA-214 suppresses growth, migration and invasion through a novel target, high mobility group AT-hook 1, in human cervical and colorectal cancer cells’

Optimisation of an Active Remote Centre of Motion Mechanism for Minimal Extracorporeal Workspace for Robotic Surgery

Conventional laparoscopic surgical tools penetrate a patient body through a fixed point called trocar point. Surgeons maintain this pivoting point on the patient body with their coordinated hand movements. Mechanism used for achieving this kinematic constraint is known as Remote Centre of Motion (RCM) mechanism and is widely used in surgical robots. Active RCM mechanism will enable positioning the constraint point virtually anywhere within the reach of the linkage. In this work, we present an optimisation strategy for an active RCM to minimise the extracorporeal workspace. The link lengths of the active RCM mechanism have been optimised and the dexterity of the tool has been analysed for various trocar positions. Kinematic analysis of the RCM mechanism, optimisation strategy, and the results of the dexterity analysis are presented.