Pelvic lymphadenectomy as a component of interval cytoreduction for ovarian cancer: is there a benefit? A pilot study

Background: Management strategy in ovarian cancer includes a combination of cytoreductive surgery and chemotherapy. Interval cytoreductive surgery has been shown to be oncologically non-inferior to primary cytoreduction with the additional benefit of reduced morbidity. Lymphadenectomy as a component of cytoreductive surgery has been controversial with an unproven therapeutic benefit.Methods: Records of patients with a histological diagnosis of ovarian cancer and treated with interval cytoreduction were evaluated. Disease related, pathological and treatment data collected for analysis.Results: The study included 32 patients with a mean age of 56 years (41-76). Serous papillary tumors (42%) were the predominate histology and the majority were in stage III disease (84%). Optimal cytoreduction was achieved in 93%. The mean nodal harvest was 9.8 nodes with left pelvic dissection yielding slightly more nodes than the right (4.5 vs 5.2). Nodal positivity was observed in just one patient (3%). A total of 314 were nodes examined with only 2 (0.6%) yielding persistent disease. The nodal positivity yield tested as a categorical variable by the binomial test returned P=0.0001.Conclusions: It is possible to omit pelvic nodal dissection during interval cytoreduction in otherwise optimally cytoreduced patients particularly when imaging and intraoperative assessment are not suggestive of pelvic nodal metastasis

Europium nanoparticle-based high performing immunoassay for the screening of treponemal antibodies

Treponema pallidum subspecies pallidum (Tp) is the causative agent of syphilis which mainly spreads through sexual contact, blood transfusion and perinatal route. In order to curtail the spread of the infection and to clinically manage the disease, timely, accurate and reliable diagnosis is very important. We have developed an immunoassay for the detection of treponemal antibodies in human serum or plasma samples. In vivo biotinylated and non-biotinylated versions of the recombinant antigen were designed by the fusion of three Tp-specific antigens namely Tp15, Tp17 and Tp47. These fusion antigens were expressed in E. coli and purified using single-step metal affinity chromatography. Biotinylated fusion antigen immobilized on streptavidin coated plate was used to capture the treponemal antibodies and the non-biotinylated antigen coated on europium nanoparticles was used as tracer. Assays with two different incubation times of 10 min and 1 h were developed, and following the incubation the europium fluorescence was measured using time-resolved fluorometry. The developed Time-resolved Fluorometric (TRF) immunoassays were evaluated with in-house and commercial serum/plasma sample panels. For well-established treponemal antibodies positive or negative samples, the sensitivity of TRF immunoassay with 10 min incubation time was 97.4% and of TRF immunoassay with 1 h incubation time was 98.7% and the specificities of both the TRF immunoassays were 99.2%. For the samples with discordant results with the reference assays, both the TRF immunoassays showed better specificity than the Enzygnost syphilis enzyme immunoassay as a screening test. The two different incubation times did not have any significant effect on the signal to cutoff (S/Co) ratios obtained with the two immunoassays (p  =  0.06). Our results indicate that the developed immunoassay with a short incubation time of 10 min has the potential to be used in clinical laboratories and in blood-bank settings as a screening test for treponemal antibodies

Escherichia coli–expressed near full length HIV-1 envelope glycoprotein is a highly sensitive and specific diagnostic antigen

Background: The Human Immunodeficiency Virus type 1 (HIV-1) envelope glycoprotein gp160, useful in detecting anti-HIV-1 antibodies, is difficult to express in heterologous hosts. The major hurdles are its signal sequence, strong hydrophobic regions and heavy glycosylation. While it has not been possible to express full length recombinant (r)-gp160 in E. coli, it can be expressed in insect and mammalian cells, but at relatively higher cost. In this work, we report E. coli-based over-expression of r-gp160 variant and evaluate its performance in diagnostic immunoassays for the detection of anti-HIV-1 antibodies. Methods: A deletion variant of r-gp160 lacking hydrophobic regions of the parent full length molecule was expressed in E. coli and purified to near homogeneity using single-step Ni (II)-affinity chromatography. Biotinylated and europium (III) chelate-labeled versions of this antigen were used to set up one- and two-step time-resolved fluorometric double antigen sandwich assays. The performance of these assays was evaluated against a collection of well-characterized human sera (n = 131), that included an in-house panel and four commercially procured panels. Results: In-frame deletion of three hydrophobic regions, spanning amino acid residues 1–43, 519–538 and 676–706, of full length HIV-1 gp160 resulted in its expression in E. coli. Both the one- and two-step assays manifested high sensitivity unambiguously identifying 75/77 and 77/77 HIV-1 positive sera, respectively. Both assays also identified all 52 HIV-seronegative sera correctly. Between the two assays, the mean signal-to-cutoff value of the two-step assay was an order of magnitude greater than that of the one-step assay. Both assays were highly specific manifesting no cross-reactivity towards antibodies specific to other viruses like hepatitis B, C and human T cell leukemia viruses. Conclusions: This study has demonstrated the expression of r-gp160 variant in E. coli, by deletion of hydrophobic regions, and its purification in reasonable yields. This underscores the potential for cost saving in antigen production. Evaluation of this antigen in a double antigen sandwich two-step assay showed it to be a highly sensitive and specific HIV-1 diagnostic reagent. The amenability of this assay to the one-step format suggests its potential utility in developing a rapid point-of-care HIV-1 diagnostic test

Retrospective analysis of random and systematic errors in radiation therapy of head and neck cancer patients and its clinical predictive implications with VMAT treatment

Background: The accuracy of radiotherapy is based on the matching of 2D portal/CBCT image with a reference image. The aim of this study is to determine the random and systematic setup errors (in cm) in radiotherapy of head and neck cancer patients and to derive the setup margin and its clinical implications.Methods: Author retrospectively reviewed the records of 25 head and neck cancer (HNC) patients treated with radiotherapy between Dec 2017 and July 2018. After immobilization, setup accuracy was assessed by registration of XVI image with planning reference image using Elekta XVI image guidance system and the isocenter correction was applied. For each patient 10 CBCT image sets were taken. The translational errors in X, Y and Z directions were used to estimate systematic (Σ) and random (σ) errors and to derive the final setup margin by using van Herk’s formula (2.5Σ + 0.7σ).Results: The mean translational errors ranges from -0.23 cm to 0.32 cm in Lateral (X), -0.15 to 0.16 cm in Longitudinal (Y) and -0.11 to 0.17 cm in vertical (Z) directions. The Mean and SD for systematic errors 0.21±0.13, 0.11±0.18, 0.14±0.11 and random error (in cm) are -0.03±0.33, 0.00±0.21 and 0.05±0.30 in X, Y and Z axis respectively. The final total margin for CTV to PTV including setup margin in the X, Y and Z directions (in cm) were 0.56, 0.61, and 0.47 respectively.Conclusion: Thus, the precise immobilization techniques are very important to reduce the setup margins, and the number of CBCTs during head and neck radiotherapy treatment

Anticonvulsant activity of gap-junctional blocker carbenoxolone in albino rats

Background: Gap junctions (GJs) are clusters of channels that connect the interiors of adjoining neurons and mediate electrical/electrotonic coupling by transfer of ions and small molecules. Electrotonic coupling between principal neurons via GJs is increasingly recognized as one of the mechanisms in the pathogenesis of the abnormal neuronal synchrony that occurs during seizures. Carbenoxolone the succinyl ester of glycyrrhetinic acid obtained from liquorice has been shown to have the property of blocking gap junctional intercellular communication. The objectives were to study if carbenoxolone has in-vivo anticonvulsive activity in pentylenetetrazole (PTZ) and maximal electroshock (MES) seizure models and to probe the functional role of GJs in seizures.Methods: Carbenoxolone was tested for anticonvulsive effect in albino rats subjected to seizures by the PTZ and MES at three doses 100 m/kg, 200 m/kg, 300 m/kg. In the PTZ model parameters observed were seizure protection, seizure latency and seizure duration. In the MES model parameters observed were seizure protection and seizure duration.Results: The results showed that the carbenoxolone has anticonvulsant activity in both PTZ and MES induced seizures with better protection in the PTZ induced seizures. In the PTZ model carbenoxolone produced a statistically significant increase in seizure latency, decrease in seizure duration and seizure protection. In the MES model carbenoxolone produced a statistically significant decrease in seizure duration.Conclusions: Carbenoxolone has in-vivo anticonvulsive effect and could be useful in both petitmal (absence) seizures and grand mal (generalized tonic-clonic epilepsy) seizures. The protective effect of carbenoxolone could be due to blockade of GJ channels that mediate electro tonic coupling and thereby prevent the neural synchronization that is characteristic of seizures. The study also supports the view that GJs have a functional role in the electrophysiology of seizures and GJ blockers have potential as a new class of antiepileptic drugs

Ultrasensitive and Robust Point-of-Care Immunoassay for the Detection of Plasmodium falciparum Malaria.

Plasmodium falciparum malaria is widespread in the tropical and subtropical regions of the world. There is ongoing effort to eliminate malaria from endemic regions, and sensitive point-of-care (POC) diagnostic tests are required to support this effort. However, current POC tests are not sufficiently sensitive to detect P. falciparum in asymptomatic individuals. After extensive optimization, we have developed a highly sensitive and robust POC test for the detection of P. falciparum infection. The test is based on upconverting nanophosphor-based lateral flow (UCNP-LF) immunoassay. The developed UCNP-LF test was validated using whole blood reference panels containing samples at different parasite densities covering eight strains of P. falciparum from different geographical areas. The limit of detection was compared to a WHO-prequalified rapid diagnostic test (RDT). The UCNP-LF achieved a detection limit of 0.2-2 parasites/μL, depending on the strain, which is 50- to 250-fold improvement in analytical sensitivity over the conventional RDTs. The developed UCNP-LF is highly stable even at 40 °C for at least 5 months. The extensively optimized UCNP-LF assay is as simple as the conventional malaria RDTs and requires 5 μL of whole blood as sample. Results can be read after 20 min from sample addition, with a simple photoluminescence reader. In the absence of a reader device at the testing site, the strips after running the test can be transported and read at a central location with access to a reader. We have found that the test and control line signals are stable for at least 10 months after running the test. The UCNP-LF has potential for diagnostic testing of both symptomatic and asymptomatic individuals

Activation of stress-related signalling pathway in human cells upon SiO2 nanoparticles exposure as an early indicator of cytotoxicity

<p>Abstract</p> <p>Background</p> <p>Nanomaterials such as SiO₂nanoparticles (SiO₂NP) are finding increasing applications in the biomedical and biotechnological fields such as disease diagnostics, imaging, drug delivery, food, cosmetics and biosensors development. Thus, a mechanistic and systematic evaluation of the potential biological and toxic effects of SiO₂NP becomes crucial in order to assess their complete safe applicability limits.</p> <p>Results</p> <p>In this study, human monocytic leukemia cell line THP-1 and human alveolar epithelial cell line A549 were exposed to a range of amorphous SiO₂NP of various sizes and concentrations (0.01, 0.1 and 0.5 mg/ml). Key biological indicators of cellular functions including cell population density, cellular morphology, membrane permeability, lysosomal mass/pH and activation of transcription factor-2 (ATF-2) were evaluated utilizing quantitative high content screening (HCS) approach and biochemical techniques. Despite the use of extremely high nanoparticle concentrations, our findings showed a low degree of cytotoxicity within the panel of SiO₂NP investigated. However, at these concentrations, we observed the onset of stress-related cellular response induced by SiO₂NP. Interestingly, cells exposed to alumina-coated SiO₂NP showed low level, and in some cases complete absence, of stress response and this was consistent up to the highest dose of 0.5 mg/ml.</p> <p>Conclusions</p> <p>The present study demonstrates and highlights the importance of subtle biological changes downstream of primary membrane and endocytosis-associated phenomena resulting from high dose SiO₂NP exposure. Increased activation of transcription factors, such as ATF-2, was quantitatively assessed as a function of i) human cell line specific stress-response, ii) SiO₂NP size and iii) concentration. Despite the low level of cytotoxicity detected for the amorphous SiO₂NP investigated, these findings prompt an in-depth focus for future SiO₂NP-cell/tissue investigations based on the combined analysis of more subtle signalling pathways associated with accumulation mechanisms, which is essential for establishing the bio-safety of existing and new nanomaterials.</p

Ultrasensitive and Robust Point-of-Care Immunoassay for the Detection of Plasmodium falciparum Malaria

Plasmodium falciparum malaria is widespread in the tropical and subtropical regions of the world. There is ongoing effort to eliminate malaria from endemic regions, and sensitive point-of-care (POC) diagnostic tests are required to support this effort. However, current POC tests are not sufficiently sensitive to detect P. falciparum in asymptomatic individuals. After extensive optimization, we have developed a highly sensitive and robust POC test for the detection of P. falciparum infection. The test is based on upconverting nanophosphor-based lateral flow (UCNP-LF) immunoassay. The developed UCNP-LF test was validated using whole blood reference panels containing samples at different parasite densities covering eight strains of P. falciparum from different geographical areas. The limit of detection was compared to a WHO-prequalified rapid diagnostic test (RDT). The UCNP-LF achieved a detection limit of 0.2-2 parasites/μL, depending on the strain, which is 50- to 250-fold improvement in analytical sensitivity over the conventional RDTs. The developed UCNP-LF is highly stable even at 40 °C for at least 5 months. The extensively optimized UCNP-LF assay is as simple as the conventional malaria RDTs and requires 5 μL of whole blood as sample. Results can be read after 20 min from sample addition, with a simple photoluminescence reader. In the absence of a reader device at the testing site, the strips after running the test can be transported and read at a central location with access to a reader. We have found that the test and control line signals are stable for at least 10 months after running the test. The UCNP-LF has potential for diagnostic testing of both symptomatic and asymptomatic individuals. </p