The potential of R. toruloides mevalonate pathway genes in increasing isoprenoid yields in S. cerevisiae: Evaluation of GGPPS and HMG-CoA reductase.

The enzymes of the mevalonate pathway need to be improved to achieve high yields of isoprenoids in the yeast Saccharomyces cerevisiae. The red yeast Rhodosporidium toruloides produces high levels of carotenoids and may have evolved to carry a naturally high flux of isoprenoids. Enzymes from such yeasts are likely to be promising candidates for improvement. Towards this end, we have systematically investigated the various enzymes of the mevalonate pathway of R. toruloides and custom synthesized, expressed, and evaluated six key enzymes in S. cerevisiae. The two nodal enzymes geranyl pyrophosphate synthase (RtGGPPS) and truncated HMG-CoA reductase (RttHMG) of R. toruloides showed a significant advantage to the cells for isoprenoid production as seen by a visual carotenoid screen. These two were analyzed further, and attempts were also made at further improvement. RtGGPPS was confirmed to be superior to the S. cerevisiae enzyme, as seen from in vitro activity determinations and in vivo production of the heterologous diterpenoid sclareol. Four mutants were created through rational mutagenesis but were unable to improve the activity further. In the case of RttHMG, functional evaluation of the enzyme revealed that it was very unstable despite functioning very well in S. cerevisiae. We succeeded in stabilizing the enzyme through mutation of a conserved serine in the catalytic region, which did not alter the enzyme activity per se. In vivo evaluation of the mutant revealed that it could enable better sclareol yields. Therefore, these two enzymes from the red yeast are excellent candidates for heterologous isoprenoid production.

A randomized trial to compare the analgesic effect of pecto-intercostal fascial plane block with erector spinae plane block after mid-sternotomy incision for cardiac surgery.

Introduction: Most cardiac surgeries are performed through a median sternotomy, of which 49% of these patients experience severe pain at rest postoperatively and up to 78% on coughing and deep breathing. Regional thoracic wall blocks targeting thoracic nerve roots improve the analgesia quality and limit opioid use. Truncal blocks through the posterior approach can often be cumbersome in patients with multiple lines and catheters. Pecto-Intercostal Fascial Plane Block (PIFB) can be a convenient alternative for achieving comparable analgesia. Material and methods: The patients were randomly assigned to receive either an ultrasound-guided Pecto-Intercostal Fascial Plane Block (PIFB) or Erector Spinae Plane Block (ESPB). The outcomes measured and compared postoperative pain scores at rest and on deep breathing at 2, 6, 12, 24 h, total opioid (fentanyl) consumption in the postoperative period, time to rescue analgesia and total rescue analgesic doses required, between the two groups. Results: Data from 30 patients were analysed. Post-operative pain scores at rest and during deep breathing were found to be comparable in both groups. The total opioid consumed, time to rescue analgesia and total doses of rescue analgesia was not found to be statistically different in the two groups. Conclusions: PIFB was found to be comparable to ESPB in alleviating post-operative pain in patients who underwent cardiac surgeries through sternotomy. And it/PIFB can be a quicker alternative to posterior truncal blocks since it can be safely given in a supine position with an ultrasound.

Incidence of postoperative pulmonary congestion as diagnosed by lung ultrasound in surgeries performed under general anaesthesia: A prospective, observational study.

Background and Aims: Administering liberal fluid raises concerns about pulmonary congestion postoperatively. Bedside ultrasonography is a valuable tool for the early detection of pulmonary congestion. In this study, we have used it to ascertain the impact of the duration of surgery and intraoperative fluid volume on the causation of pulmonary congestion. Our objective was to determine the incidence of pulmonary congestion as diagnosed by lung ultrasound in patients undergoing general anaesthesia with varied fluid administration. Methods: Seventy participants of American Society of Anesthesiologists physical status I and II, aged between 18 and 60 years, undergoing elective extrathoracic surgeries of over 3 h under general anaesthesia were included. Preoperative lung ultrasound was carried out in all patients, and a postoperative lung ultrasound was carried out at 1 h after extubation. The appearance of three or more "B"-lines was considered positive for lung congestion. Results: Significant differences (P < 0.001) were found in the duration of surgery and the appearance of B-lines in the postoperative period. Participants who developed B lines received, on average, 150% more fluid (1148.16 ± 291.79 ml) than those who did not (591.29 ± 398.42 ml) (P = 0.0240). Net fluid balance was also significantly different in patients who developed B lines (P = 0.0014). None of the patients developed symptoms of lung congestion postoperatively. Conclusion: Long duration of surgery under general anaesthesia (>3 h) with the administration of large volumes of intraoperative fluid and a large net fluid balance are associated with lung congestion as diagnosed by lung ultrasound.

Mode of Mechanical Ventilation in a Case of Venolymphatic Malformation: Spontaneous-Saves, Positive-Precludes.

Mediastinal venolymphatic malformations (VLM) are rare tumours, with very few reported cases in the literature. Arising often from the anterior mediastinum, VLM manifests symptoms based on invaded surrounding structures. Masses from the anterior and superior mediastinum pose an anaesthetic challenge for airway and hemodynamic management. A 7-month-old male child presented with a progressively growing mass over the left anterior chest wall for one month, about 4x4 cm, with diffuse margins and now expanded to involve the root of the neck and into the axilla. The patient was free from any apparent systemic illness. The breathing difficulty worsened in the past week with noisy respiration associated with feeding difficulty and hence sought medical admission to the paediatrics emergency unit. In conclusion, such huge mediastinal masses are managed better under spontaneous ventilation with an adequate surgical depth of anaesthesia to maintain appropriate respiratory compliance and necessitate lower peak inspiratory pressure. Given rare cases reported in the literature, similar topics would help choose the modus of ventilation and their safe management.

Role of Indocyanine Green Angiography in Free Flap Surgery: A Comparative Outcome Analysis of a Single-Center Large Series of 877 Consecutive Free Flaps.

Purpose This study aims to assess and validate the role and cost-effectiveness of indocyanine green angiography (ICGA) in free flap surgery outcomes. A new intraoperative protocol of whole-body surface warming (WBSW) for all free flap surgeries during the strategic "microbreaks" is also described. Methods A retrospective analysis of 877 consecutive free flaps, performed over 12 years, is presented. The results of the ICGA group ( n = 438) were compared with the historical No-ICGA group ( n = 439), and statistical significance was calculated for three crucial flap-related adverse outcomes and cost-effectiveness. ICGA was also used as a tool to show the effect of WBSW on free flaps. Results ICGA showed a notably strong statistical significance in decreasing two outcome parameters, namely, partial flap loss and re-exploration rate. It was also cost-effective. ICGA also demonstrated the positive role of WBSW in increasing flap perfusion. Conclusions Our study shows that the usage of ICGA for intraoperative assessment of flap perfusion can significantly reduce the partial flap loss and re-exploration rate in free flap surgeries in a cost-effective manner. A new protocol of WBSW is also described and recommended to increase flap perfusion in all free flap surgeries.

First Case Report of Breast Implant Associated-Anaplastic Large Cell Lymphoma from India: Are We Ready?

Breast implant associated-anaplastic large cell lymphoma (BIA-ALCL) has become a hot topic in recent plastic surgery and oncology forum. Its cases have been on the rise since its first emergence more than two decades ago. This condition is less known and management guidelines are still evolving. BIA-ALCL was seen recently with a classical presentation in one of our patients, who underwent immediate reconstruction with a macro-textured silicone implant following breast cancer surgery. We want to add the first case report from India to the global information database. There are still unanswered questions in its management, and we wish to highlight the same to make way for further research. With the rise in aesthetic and reconstructive implant surgeries, the knowledge of BIA-ALCL must expand among oncologists, radiologists, and pathologists for early identification and treatment for better patient outcomes.

Transillumination and Point-of-Care Ultrasonography to Delineate Tracheal Deviation for Challenging Tracheostomy: A Case Report.

Tracheostomy is indicated for varied reasons and is a relatively safe procedure that can be done with both open and percutaneous methods. However, the procedure is often challenging in cases of distorted neck anatomy. Neck swellings often push the trachea laterally or shield it. Even some gross intrathoracic pathology may shift the trachea from the typical trajectory making it challenging to delineate the course of the trachea. A bedside point-of-care technique having a visual aid that can guide the performer thus appears beneficiary. Fiber-optic assistance for correct puncture and confirmation is known, and light-based techniques have been used for tracheostomies. As fiberscopes are not infrequent in tertiary and even secondary care hospitals, transillumination from a flexible bronchoscope can identify the altered course of the trachea, much like a navigation system, and systematically aid the performer in steering away from the obstacles. We present two cases in two scenarios with tracheal deviation who underwent either open or percutaneous tracheostomy with point-of-care ultrasound and transillumination to delineate the course of the trachea and facilitate difficult tracheostomies safely.

Efficacy of flupirtine for postoperative pain: A systematic review and meta-analysis.

Background and Aims: Flupirtine has been used for various chronic pain conditions, but its utility in the perioperative period as an analgesic is still inconclusive. This systematic review and meta-analysis aimed to assess the efficacy of flupirtine for postoperative pain. Methods: PubMed, Embase and the Cochrane Central Register of Controlled Trials (CENTRAL) were explored for the randomised controlled trials (RCTs) which compared flupirtine with other analgesic/placebo for perioperative pain in adult patients undergoing surgery. The standardised mean difference (SMD) of pain scores, the need for rescue analgesia and all adverse effects were assessed. Heterogeneity was assessed using Cochrane's Q statistic test and I2 statistic. Cochrane Collaboration's tool was used to evaluate the risk of bias and the quality of the RCTs. Results: A total of 13 RCTs (including 1,014 patients) that evaluated the use of flupirtine for postoperative pain were included in the study. The pooled SMD of postoperative pain scores revealed that flupirtine and other analgesics were comparable at 0, 6, 12 and 24 hours (P > 0.05), while at 48 hours, flupirtine showed poor pain control (P = 0.04) as compared to other analgesics. There were no significant differences at other time points and on comparison of flupirtine with placebo. The side effect profile was comparable between flupirtine and other analgesics. Conclusion: The current evidence suggests that perioperative flupirtine was not superior to other most commonly used analgesics and placebo for the treatment of postoperative pain.

Transcriptomic data meta-analysis reveals common and injury model specific gene expression changes in the regenerating zebrafish heart.

Zebrafish have the capacity to fully regenerate the heart after an injury, which lies in sharp contrast to the irreversible loss of cardiomyocytes after a myocardial infarction in humans. Transcriptomics analysis has contributed to dissect underlying signaling pathways and gene regulatory networks in the zebrafish heart regeneration process. This process has been studied in response to different types of injuries namely: ventricular resection, ventricular cryoinjury, and genetic ablation of cardiomyocytes. However, there exists no database to compare injury specific and core cardiac regeneration responses. Here, we present a meta-analysis of transcriptomic data of regenerating zebrafish hearts in response to these three injury models at 7 days post injury (7dpi). We reanalyzed 36 samples and analyzed the differentially expressed genes (DEG) followed by downstream Gene Ontology Biological Processes (GO:BP) analysis. We found that the three injury models share a common core of DEG encompassing genes involved in cell proliferation, the Wnt signaling pathway and genes that are enriched in fibroblasts. We also found injury-specific gene signatures for resection and genetic ablation, and to a lower extent the cryoinjury model. Finally, we present our data in a user-friendly web interface that displays gene expression signatures across different injury types and highlights the importance to consider injury-specific gene regulatory networks when interpreting the results related to cardiac regeneration in the zebrafish. The analysis is freely available at: https://mybinder.org/v2/gh/MercaderLabAnatomy/PUB_Botos_et_al_2022_shinyapp_binder/HEAD?urlpath=shiny/bus-dashboard/ .

Wt1 transcription factor impairs cardiomyocyte specification and drives a phenotypic switch from myocardium to epicardium.

During development, the heart grows by addition of progenitor cells to the poles of the primordial heart tube. In the zebrafish, Wilms tumor 1 transcription factor a (wt1a) and b (wt1b) genes are expressed in the pericardium, at the venous pole of the heart. From this pericardial layer, the proepicardium emerges. Proepicardial cells are subsequently transferred to the myocardial surface and form the epicardium, covering the myocardium. We found that while wt1a and wt1b expression is maintained in proepicardial cells, it is downregulated in pericardial cells that contributes cardiomyocytes to the developing heart. Sustained wt1b expression in cardiomyocytes reduced chromatin accessibility of specific genomic loci. Strikingly, a subset of wt1a- and wt1b-expressing cardiomyocytes changed their cell-adhesion properties, delaminated from the myocardium and upregulated epicardial gene expression. Thus, wt1a and wt1b act as a break for cardiomyocyte differentiation, and ectopic wt1a and wt1b expression in cardiomyocytes can lead to their transdifferentiation into epicardial-like cells.

Utility and futility of central venous catheterization.

Central venous access is useful for monitoring central venous pressure, inserting pulmonary artery catheter and administering vasoactive drugs in hemodynamically unstable patients. Central venous catheter (CVC) insertion through internal jugular vein may cause major vessel injury, inadvertent arterial catheterization, brachial plexus injury, phrenic nerve injury, pneumothorax, and haemothorax. We describe unusual presentation of hemothorax following CVC placement in a patient undergoing vestibular schwannoma excision. The patients' trachea intubated after several attempts during which thiopentone up to 600 mg administered. Thereafter, under ultrasound guidance, an 18G introducer needle placed in the right internal jugular vein but guide-wire did not advance. Meanwhile, the patient became hemodynamically unstable and a CVC placed in right subclavian vein and norepinephrine infused at 0.05 µg/kg/min; simultaneously, 1000 ml normal saline administered through CVC. The hemodynamic instability attributed to thiopentone administered during endotracheal intubation. The surgical procedure cancelled, and the patient shifted to critical care unit (CCU). Mechanical ventilation continued. In CCU, hemodynamic parameters further deteriorated and 0.1 µg/kg/min epinephrine started. Bedside lung ultrasound showed a large collection in pleural space on the right side. Chest radiograph showed a homogenous opacity obliterating costophrenic angle on the right side. A possibility of hemothorax considered, chest tube inserted and 1000 ml sanguineous fluid drained. Blood sample drawn through CVC showed air from proximal and middle lumen but distal lumen drained blood. Another CVC placed in the femoral vein and subclavian vein CVC removed. The vasoactive drug infusion transferred to CVC in femoral vein and 2 units pRBCs transfused. Hemodynamic parameters gradually stabilized and the patient recovered completely.

New 'ZISIS' (Zigzag Incision-Suction-Irrigation System) Technique for Endoscopic-Assisted Forehead Tumour Excision.

BACKGROUND: Endoscopic-assisted excision of forehead tumours like osteomas and lipomas is well established, but the conventional techniques suffer from many limitations like inadequate access, fogging of the endoscope and unclear vision due to collection of blood and debris. METHOD: Three simple modifications of the conventional endoscopic forehead technique for benign tumour excisions are described by the acronym 'ZISIS'. ZI Zigzag scalp incision increases the surface area of the opening permitting easy insertion of multiple instruments along with the endoscope.S Suction tubing made from a disposable suction catheter tube is taped along the endoscope sheath for continuous suctioning and good vision.IS Irrigation system of warm saline made with an infant feeding tube is also taped along the endoscope just opening in the front of the suction tube. RESULTS: A total of 12 consecutive patients underwent endoscopic excision of forehead benign tumours in 2 years. This included 2 lipomas and 10 osteomas cases. All patients achieved excellent hidden scars in the scalp. All patients rated their results as excellent with respect to the hidden scar and aesthetic result. No early or late complications were reported with follow-up ranging from 6 months to 2 years. CONCLUSION: A new simple modification called 'ZISIS' endoscopic forehead excision technique is described for benign forehead tumours excision making it easier, efficient and ergonomic. LEVEL OF EVIDENCE IV: Evidence obtained from multiple time series with or without the intervention, such as case studies.

A new tension induction paradigm unravels tissue response and the importance of E-cadherin in the developing epidermis.

The epidermis, being the outermost epithelial layer in metazoans, experiences multiple external and self-generated mechanical stimuli. The tissue-scale response to these mechanical stresses has been actively studied in the adult stratified epidermis. However, the response of the developing bi-layered epidermis to differential tension and its molecular regulation has remained poorly characterised. Here we report an oil injection based method, which in combination with atomic force microscopy (AFM), allows manipulation as well as estimation of tension in the developing epidermis. Our results show that the injection of mineral oil into the brain ventricle of developing zebrafish embryos stretches the overlying epidermis. The epidermal tension increases linearly with the injected volume of oil and the injection of 14-17 nL oil results in a two-fold increase in epidermal tension. This increase in epidermal tension is sufficient to elicit a physiological response characterised by temporal changes in the cell cross-sectional area and an increase in cell proliferation. Our data further indicate that the depletion of E-cadherin in the epidermis is detrimental for tissue integrity under increased mechanical stress. The application of this experimental paradigm in a genetically tractable organism such as zebrafish can be useful in uncovering mechanisms of tension sustenance in the developing epidermis.