Impact of non-invasive post-mortem micro-CT imaging on a fetal autopsy service: a single centre retrospective study.

AIM: To evaluate the impact of a new, less-invasive micro-computed tomography (CT) service on autopsy service provision. We recorded parental consent, type of autopsy performed, autopsy reporting times and time taken for the body to be released from the mortuary. MATERIALS AND METHODS: A retrospective, single-centre case series was conducted for all perinatal deaths since the introduction of our micro-CT service in 2016, with a detailed review of records extracted from 2019 and 2021. Fetal demographics (gestational age, weight), type of autopsy conducted, and the time taken from receiving the body to releasing the body and issuing a final report were recorded. RESULTS: Micro-CT imaging uptake increased to over two hundred cases/year by 2021. Overall, invasive autopsies reduced from (45.8%, 196/428; 2019) to (32.1%, 125/390; 2021) with an equivalent rise in less-invasive autopsy from 54.2% (232/428;2019) to 67.9% (265/390;2019). Offering a micro-CT service resulted in an increase in consent to imaging-based autopsies from (76.9%, 329/428;2019) to (87.2%, 340/390;2021). Micro-CT has become the most common post-mortem imaging performed in our institution at 54.4% (212/251;2021), although the body preparation time from the tissue staining required has increased the time to provide an autopsy report to 17 days and release of the body to 18 days. CONCLUSION: Our study shows that introducing a micro-CT post-mortem imaging service was associated with reduced use of conventional invasive procedures, despite a slight increase in turnaround times. Understanding these factors and continued improvements in micro-CT service delivery will help make this accessible to a wider population in the future.

A Priori Activation of Apoptosis Pathways of Tumor (AAAPT) technology: Development of targeted apoptosis initiators for cancer treatment.

Cancer cells develop tactics to circumvent the interventions by desensitizing themselves to interventions. Amongst many, the principle routes of desensitization include a) activation of survival pathways (e.g. NF-kB, PARP) and b) downregulation of cell death pathways (e.g. CD95/CD95L). As a result, it requires high therapeutic dose to achieve tumor regression which, in turn damages normal cells through the collateral effects. Methods are needed to sensitize the low and non-responsive resistant tumor cells including cancer stem cells (CSCs) in order to evoke a better response from the current treatments. Current treatments including chemotherapy can induce cell death only in bulk cancer cells sparing CSCs and cancer resistant cells (CRCs) which are shown to be responsible for high recurrence of disease and low patient survival. Here, we report several novel tumor targeted sensitizers derived from the natural Vitamin E analogue (AMP-001-003). The drug design is based on a novel concept "A priori activation of apoptosis pathways of tumor technology (AAAPT) which is designed to activate specific cell death pathways and inhibit survival pathways simultaneously and selectively in cancer cells sparing normal cells. Our results indicate that AMP-001-003 sensitize various types of cancer cells including MDA-MB-231 (triple negative breast cancer), PC3 (prostate cancer) and A543 (lung cancer) cells resulting in reducing the IC-50 of doxorubicin in vitro when used as a combination. At higher doses, AMP-001 acts as an anti-tumor agent on its own. The synergy between AMP-001 and doxorubicin could pave a new pathway to use AAAPT leading molecules as neoadjuvant to chemotherapy to achieve better efficacy and reduced off-target toxicity compared to the current treatments.

Feasibility of INTACT (INcisionless TArgeted Core Tissue) biopsy procedure for perinatal autopsy.

OBJECTIVES: To determine the feasibility and tissue yield of a perinatal incisionless ultrasound-guided biopsy procedure, the INcisionless Targeted Core Tissue (INTACT) technique, in the context of minimally invasive autopsy. METHODS: Cases of perinatal death in which the parents consented for minimally invasive autopsy underwent postmortem magnetic resonance imaging and an INTACT biopsy procedure, defined as needle biopsy of organs via the umbilical cord, performed under ultrasound guidance. In each case, three cores of tissue were obtained from seven target organs (both lungs, both kidneys, heart, spleen and liver). Biopsy success was predefined as an adequate volume of the intended target organ for pathological analysis, as judged by a pathologist blinded to the case and biopsy procedure. RESULTS: Thirty fetuses underwent organ sampling. Mean gestational age was 30 weeks (range, 18-40 weeks) and mean delivery-to-biopsy interval was 12 days (range, 6-22 days). The overall biopsy success rate was 153/201 (76.1%) samples, with the success rates in individual organs being highest for the heart and lungs (93% and 91%, respectively) and lowest for the spleen (11%). Excluding splenic samples, the biopsy success rate was 150/173 (86.7%). Histological abnormalities were found in 4/201 (2%) samples, all of which occurred in the lungs and kidneys of a fetus with pulmonary hypoplasia and multicystic kidney disease. CONCLUSIONS: Incisionless ultrasound-guided organ biopsy using the INTACT procedure is feasible, with an overall biopsy success rate of over 75%. This novel technique offers the ideal combination of an imaging-led autopsy with organ sampling for parents who decline the conventional invasive approach. © 2019 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of the International Society of Ultrasound in Obstetrics and Gynecology.

Molecular fingerprinting of the Indian medicinal plant Strychnos minor Dennst.

Genomic DNA of Strychnos minor (Dennst) of sixteen geographical regions were amplified using thirteen primers. The analyses of products of Radom Amplified Polymorphic DNA (RAPD) revealed polymorphism among the samples. The polymorphic information content (PIC) was maximum for the primer of OPB-04 (0.40) followed by the primer OPB-20 (0.39). Clustering analysis by Jaccard's coefficient index exhibited similarity matrix and distance matrix. The amplified products of the 13 primers collectively showed similarity index ranged from 0.654 to 0.986. The phylogenetic trees were constructed by UPGMA cluster analysis. Significant variations in the amplified genomic DNA suggest genetic variability of population but low inter-population genetic segregation.

Optimization of Processing Parameters in ECM of Die Tool Steel Using Nanofluid by Multiobjective Genetic Algorithm.

Formation of spikes prevents achievement of the better material removal rate (MRR) and surface finish while using plain NaNO3 aqueous electrolyte in electrochemical machining (ECM) of die tool steel. Hence this research work attempts to minimize the formation of spikes in the selected workpiece of high carbon high chromium die tool steel using copper nanoparticles suspended in NaNO3 aqueous electrolyte, that is, nanofluid. The selected influencing parameters are applied voltage and electrolyte discharge rate with three levels and tool feed rate with four levels. Thirty-six experiments were designed using Design Expert 7.0 software and optimization was done using multiobjective genetic algorithm (MOGA). This tool identified the best possible combination for achieving the better MRR and surface roughness. The results reveal that voltage of 18 V, tool feed rate of 0.54 mm/min, and nanofluid discharge rate of 12 lit/min would be the optimum values in ECM of HCHCr die tool steel. For checking the optimality obtained from the MOGA in MATLAB software, the maximum MRR of 375.78277 mm(3)/min and respective surface roughness Ra of 2.339779 µm were predicted at applied voltage of 17.688986 V, tool feed rate of 0.5399705 mm/min, and nanofluid discharge rate of 11.998816 lit/min. Confirmatory tests showed that the actual performance at the optimum conditions was 361.214 mm(3)/min and 2.41 µm; the deviation from the predicted performance is less than 4% which proves the composite desirability of the developed models.

Proteomic identification of pterostilbene-mediated anticancer activities in HepG2 cells.

In the present study, we attempt to shed light on the underlying molecular mechanism of the anticancer activity of pterostilbene (PTS) in HepG2 cells through the proteomic approach. PTS was found to induce apoptosis by altering the expression of apoptotic genes and the G2/M phase of cell cycle arrest. Further, the 2-DE map showed the expression of 72 differentially regulated proteins in PTS-treated HepG2 cells, of which 8 spots with >2 fold up- or down-regulated level were identified by MALDI-TOF analysis, which has a regulatory role in apoptosis. These findings for the first time offer valuable insights into the mechanism of apoptotis by PTS in HepG2 cells.

The impact of oxidative stress on islet transplantation and monitoring the graft survival by non-invasive imaging.

Islet transplantation is an attractive strategy to treat severe diabetic conditions in patients suffering from autoimmune derived diabetes, and it has currently been considered a forefront research arena in diabetes. Major aim of islet transplantation is to achieve successful insulin independent disease free survival. The key challenges in transplanted islets are the generation of reactive oxygen species (ROS) and associated oxidative stress, pro-inflammatory cytokine - (TNFα) mediated apoptotic induction, attack by immune cells, and achieving revascularization with minimal hypoxic microenvironment. Free radicals and their derivatives are constantly produced in living systems, but at relatively low level, and in a balanced state. Oxidative stress, which occurs as a result of an imbalance between the intracellular free radicals production and the cellular antioxidant defense mechanisms in the transplanted islets, can lead to cell death. The balance between oxidants and antioxidants in a cell can be easily disturbed by increase in ROS production or reduction in the level of cellular antioxidant defensive substances, which can cause many metabolic complications, including pancreatic ß-cell damage. Antioxidants function as blockers of radical processes by eliminating harmful ROS produced during normal cellular metabolism. A complex antioxidant defense mechanism has been developed by nature in cells to protect the cellular homeostasis. This system mainly includes antioxidant enzymes, vitamins and minerals. As transplanted islet survival is crucial for achieving successful therapy, most of these antioxidants can be used as a supplement to scavenge the local ROS thereby improving the survival of transplanted islets. Currently, very few techniques have been routinely used to qualitatively and quantitatively assess the survival and function of islet grafts, especially to confirm the success of treatment, which includes metabolic parameters such as blood glucose, insulin and C-peptide levels. These biochemical measurements provide markers at only the late stages of islet rejection. Use of molecular imaging techniques has the potential for real-time non-invasive monitoring of the functional status and viability of transplanted islet grafts in living animals. This review mainly focuses on the current status of islet transplantations, potential preventive strategies used to reduce oxidative stress-mediated toxicity in islet grafts, and use of molecular imaging as a tool to quantitatively evaluate the functional status of the transplanted islets in living animals.

Dual-therapeutic reporter genes fusion for enhanced cancer gene therapy and imaging.

Two of the successful gene-directed enzyme prodrug therapies include herpes simplex virus-thymidine kinase (HSV1-TK) enzyme-ganciclovir prodrug and the Escherichia coli nitroreductase (NTR) enzyme-CB1954 prodrug strategies; these enzyme-prodrug combinations produce activated cytotoxic metabolites of the prodrugs capable of tumor cell death by inhibiting DNA synthesis and killing quiescent cells, respectively. Both these strategies also affect significant bystander cell killing of neighboring tumor cells that do not express these enzymes. We have developed a dual-combination gene strategy, where we identified HSV1-TK and NTR fused in a particular orientation can effectively kill tumor cells when the tumor cells are treated with a fusion HSV1-TK-NTR gene- along with a prodrug combination of GCV and CB1954. In order to determine whether the dual-system demonstrate superior therapeutic efficacy than either HSV1-TK or NTR systems alone, we conducted both in vitro and in vivo tumor xenograft studies using triple negative SUM159 breast cancer cells, by evaluating the efficacy of cell death by apoptosis and necrosis upon treatment with the dual HSV1-TK genes-GCV-CB1954 prodrugs system, and compared the efficiency to HSV1-TK-GCV and NTR-CB1954. Our cell-based studies, tumor regression studies in xenograft mice, histological analyses of treated tumors and bystander studies indicate that the dual HSV1-TK-NTR-prodrug system is two times more efficient even with half the doses of both prodrugs than the respective single gene-prodrug system, as evidenced by enhanced apoptosis and necrosis of tumor cells in vitro in culture and xenograft of tumor tissues in animals.

Noninvasive optical imaging of nitroreductase gene-directed enzyme prodrug therapy system in living animals.

Gene-directed enzyme prodrug therapy (GDEPT) is a promising and emerging strategy that attempts to limit the systemic toxicity inherent to cancer chemotherapy by means of tumor-targeted delivery and expression of an exogenous gene whose product converts nontoxic prodrug(s) into activated cytotoxic agent(s). The bacterial nitroreductase (NTR) enzyme, coupled with its substrate prodrug 5-(azaridin-1-yl)-2,4-dinitrobenzamide (CB1954), is a promising GDEPT strategy that has reached clinical trials. However, no strategy exists to visually monitor and quantitatively evaluate the therapeutic efficacy of NTR/CB1954 prodrug therapy in cells and imaging in living animals. As the success of any GDEPT is dependent upon the efficiency of transgene expression in vivo, we developed a safe, sensitive and reproducible noninvasive imaging method to monitor NTR transgene expression that would allow quantitative assessment of both therapeutic efficacy and diagnostic outcome of NTR/CB1954 prodrug therapy in the future. Here, we investigate the use of a novel fluorescent imaging dye CytoCy5S (a Cy5-labeled quenched substrate of NTR enzyme) on various cancer cell lines in vitro and in NTR-transfected tumor-bearing animals in vivo. CytoCy5S-labeled cells become fluorescent at 'red-shifted' wavelengths (638 nm) when reduced by cellular NTR enzyme and remains trapped within the cells for extended periods of time. The conversion and entrapment was dynamically recorded using a time-lapsed microscopy. Systemic and intratumoral delivery of CytoCy5S to NTR-expressing tumors in animals indicated steady and reproducible signals even 16 h after delivery (P<0.001). This is the first study to address visual monitoring and quantitative evaluation of NTR activity in small animals using CytoCy5S, and establishes the capability of NTR to function as an imageable reporter gene.

Pulmonary functions during peritoneal dialysis.

Lung volumes, flows, flow volume loop, diffusing capacity and closing volume were studied in 7 patients (Group I) before, with 2 liter dialyzate in the peritoneal cavity and after 8 hours of peritoneal dialysis. The functional residual capacity, residual volume and the total lung capacity decreased (P less than 0.01, less than 0.05, and less than 0.05 respectively) with 2 liter dialyzate in the peritoneal cavity and returned to baseline after the dialyzate was removed. Arterial blood gas analysis was performed in another 8 patients (Group II) before, with 2 liter dialyzate in the peritoneal cavity, and after the dialyzate had been drained. No significant difference was observed in PaO2, PaCO2 or pHa during any phase of the study. It is concluded that except for a minor decrease in FRC, RV and TLC, there were no deleterious effects with the use of 2 liter dialyzate volume on gas exchange or pulmonary functions during peritoneal dialysis.

Extracorporeal membrane oxygenator support in a case of severe status asthmaticus.

Support with extracorporeal membrane oxygenation (ECMO) can sustain life for a substantial period of time, but the role of long-term ECMO support is limited. The National Institutes of Health ECMO collaborative study revealed no significant reduction in mortality in adults with severe respiratory distress syndrome treated with ECMO. The value of ECMO in the treatment of transient, severe acute respiratory insufficiency has been demonstrated in patients after cardiac operations, in patients with Pneumocystis carinii pneumonia, and in patients following trauma. We present an example of refractory respiratory failure in a young asthmatic with massive atelectasis in whom the institution of ECMO allowed us to perform adequate pulmonary lavage. It resulted in improvement in gas exchange and the patient's survival.

Survival after prolonged submersion in cold water without neurologic sequelae. Report of two cases.

We describe two patients who suffered prolonged (greater than four minutes) submersion in cold water (25 minutes, 32 s, and six minutes, respectively), with complete neurologic recovery. We postulate that hypothermia served to protect the brain from hypoxic injury. These cases also serve to emphasize the need for vigorous one-the-scene resuscitative efforts. We review the problems associated with the treatment of hypothermia and drowning.

The effect of isoproterenol on hypoxia-mediated histamine release.

In 13 guinea pigs the pulmonary circulation was isolated from the systemic circulation and perfused with Tris buffer alone or with isoproterenol. The lungs were initially ventilated with 100% O2 and then switched to 100% N2 to induce alveolar hypoxia. The effluent was collected from the aorta and assayed for histamine. In the control group (n = 7) the histamine levels rose markedly with hypoxia. In the group perfused with Tris buffer and isoproterenol (n = 6) the rise in histamine levels was markedly diminished. Therefore, the authors conclude that isoproterenol, in their in vivo model, had a dramatic effect on hypoxia (non-immunologic) mediated histamine release.