Activation of protein kinase C and protein kinase D in human natural killer cells: effects of tributyltin, dibutyltin, and tetrabromobisphenol A

Up to now, the ability of target cells to activate protein kinase C (PKC) and protein kinase D (PKD) (which is often a downstream target of PKC) has not been examined in natural killer (NK) lymphocytes. Here we examined whether exposure of human NK cells to lysis sensitive tumor cells activated PKC and PKD. The results of these studies show for the first time that activation of PKC and PKD occurs in response to target cell binding to NK cells. Exposure of NK cells to K562 tumor cells for 10 and 30 min increased phosphorylation/activation of both PKC and PKD by roughly 2-fold. Butyltins (tributyltin (TBT), dibutyltin (DBT)) and brominated compounds (tetrabromobisphenol A (TBBPA)) are environmental contaminants that are found in human blood. Exposures of NK cells to TBT, DBT, or TBBPA decrease NK cell lytic function in part by activating the mitogen-activated protein kinases (MAPKs) that are part of the NK lytic pathway. We established that PKC and PKD are part of the lytic pathway upstream of MAPKs and thus we investigated whether DBT, TBT, and TBBPA exposures activated PKC and PKD. TBT-activated PKC by 2–3-folds at 10 min at concentrations ranging from 50 to 300 nM while DBT caused a 1.3-fold activation at 2.5 µM at 10 min. Both TBT and DBT caused an approximately 2-fold increase in phosphorylation/activation of PKC. Exposures to TBBPA caused no statistically significant changes in either PKC or PKD activation

Role of Protein Kinase C in TBT-Induced Inhibition of Lytic Function and MAPK Activation in Human Natural Killer Cells

Human natural killer (NK) cells are lymphocytes that destroy tumor and virally infected cells. Previous studies have shown that exposure of NK cells to tributyltin (TBT) greatly diminishes their ability to destroy tumor cells (lytic function) while activating mitogen-activated protein kinases (MAPK) (p44/42, p38, and JNK) in NK cells. The signaling pathway that regulates NK lytic function appears to include activation of protein kinase C (PKC) as well as MAPK activity. TBT-induced activation of MAPKs would trigger a portion of the NK lytic signaling pathway, which would then leave the NK cell unable to trigger this pathway in response to a subsequent encounter with a target cell. In the present study we evaluated the involvement of PKC in inhibition of NK lysis of tumor cells and activation of MAPKs caused by TBT exposure. TBT caused a 2–3-fold activation of PKC at concentrations ranging from 50 to 300 nM (16–98 ng/ml), indicating that activation of PKC occurs in response to TBT exposure. This would then leave the NK cell unable to respond to targets. Treatment with the PKC inhibitor, bisindolylmaleimide I, caused an 85% decrease in the ability of NK cells to lyse tumor cells, validating the involvement of PKC in the lytic signaling pathway. The role of PKC in the activation of MAPKs by TBT was also investigated using bisindolylmaleimide I. The results indicated that, in NK cells where PKC activation was blocked, there was no activation of the MAPK, p44/42 in response to TBT. However, TBT-induced activation of the MAPKs, p38 and JNK did not require PKC activation. These results indicate the pivotal role of PKC in the TBT-induced loss of NK lytic function including activation of p44/42 by TBT in NK cells

In Vitro Control of Post-Harvest Fruit Rot Fungi by Some Plant Essential Oil Components

Eight substances that are main components of the essential oils from three Mediterranean aromatic plants (Verbena officinalis, Thymus vulgaris and Origanum vulgare), previously found active against some phytopathogenic Fungi and Stramenopila, have been tested in vitro against five etiological agents of post-harvest fruit decay, Botrytis cinerea, Penicillium italicum, P. expansum, Phytophthora citrophthora and Rhizopus stolonifer. The tested compounds were β-fellandrene, β-pinene, camphene, carvacrol, citral, o-cymene, γ-terpinene and thymol. Citral exhibited a fungicidal action against P. citrophthora; carvacrol and thymol showed a fungistatic activity against P. citrophthora and R. stolonifer. Citral and carvacrol at 250 ppm, and thymol at 150 and 250 ppm stopped the growth of B. cinerea. Moreover, thymol showed fungistatic and fungicidal action against P. italicum. Finally, the mycelium growth of P. expansum was inhibited in the presence of 250 ppm of thymol and carvacrol. These results represent an important step toward the goal to use some essential oils or their components as natural preservatives for fruits and foodstuffs, due to their safety for consumer healthy and positive effect on shelf life extension of agricultural fresh products

The impact of immediate breast reconstruction on the time to delivery of adjuvant therapy: the iBRA-2 study

Background: Immediate breast reconstruction (IBR) is routinely offered to improve quality-of-life for women requiring mastectomy, but there are concerns that more complex surgery may delay adjuvant oncological treatments and compromise long-term outcomes. High-quality evidence is lacking. The iBRA-2 study aimed to investigate the impact of IBR on time to adjuvant therapy. Methods: Consecutive women undergoing mastectomy ± IBR for breast cancer July–December, 2016 were included. Patient demographics, operative, oncological and complication data were collected. Time from last definitive cancer surgery to first adjuvant treatment for patients undergoing mastectomy ± IBR were compared and risk factors associated with delays explored. Results: A total of 2540 patients were recruited from 76 centres; 1008 (39.7%) underwent IBR (implant-only [n = 675, 26.6%]; pedicled flaps [n = 105,4.1%] and free-flaps [n = 228, 8.9%]). Complications requiring re-admission or re-operation were significantly more common in patients undergoing IBR than those receiving mastectomy. Adjuvant chemotherapy or radiotherapy was required by 1235 (48.6%) patients. No clinically significant differences were seen in time to adjuvant therapy between patient groups but major complications irrespective of surgery received were significantly associated with treatment delays. Conclusions: IBR does not result in clinically significant delays to adjuvant therapy, but post-operative complications are associated with treatment delays. Strategies to minimise complications, including careful patient selection, are required to improve outcomes for patients

Co-infection and ICU-acquired infection in COIVD-19 ICU patients: a secondary analysis of the UNITE-COVID data set

Background: The COVID-19 pandemic presented major challenges for critical care facilities worldwide. Infections which develop alongside or subsequent to viral pneumonitis are a challenge under sporadic and pandemic conditions; however, data have suggested that patterns of these differ between COVID-19 and other viral pneumonitides. This secondary analysis aimed to explore patterns of co-infection and intensive care unit-acquired infections (ICU-AI) and the relationship to use of corticosteroids in a large, international cohort of critically ill COVID-19 patients.Methods: This is a multicenter, international, observational study, including adult patients with PCR-confirmed COVID-19 diagnosis admitted to ICUs at the peak of wave one of COVID-19 (February 15th to May 15th, 2020). Data collected included investigator-assessed co-infection at ICU admission, infection acquired in ICU, infection with multi-drug resistant organisms (MDRO) and antibiotic use. Frequencies were compared by Pearson's Chi-squared and continuous variables by Mann-Whitney U test. Propensity score matching for variables associated with ICU-acquired infection was undertaken using R library MatchIT using the "full" matching method.Results: Data were available from 4994 patients. Bacterial co-infection at admission was detected in 716 patients (14%), whilst 85% of patients received antibiotics at that stage. ICU-AI developed in 2715 (54%). The most common ICU-AI was bacterial pneumonia (44% of infections), whilst 9% of patients developed fungal pneumonia; 25% of infections involved MDRO. Patients developing infections in ICU had greater antimicrobial exposure than those without such infections. Incident density (ICU-AI per 1000 ICU days) was in considerable excess of reports from pre-pandemic surveillance. Corticosteroid use was heterogenous between ICUs. In univariate analysis, 58% of patients receiving corticosteroids and 43% of those not receiving steroids developed ICU-AI. Adjusting for potential confounders in the propensity-matched cohort, 71% of patients receiving corticosteroids developed ICU-AI vs 52% of those not receiving corticosteroids. Duration of corticosteroid therapy was also associated with development of ICU-AI and infection with an MDRO.Conclusions: In patients with severe COVID-19 in the first wave, co-infection at admission to ICU was relatively rare but antibiotic use was in substantial excess to that indication. ICU-AI were common and were significantly associated with use of corticosteroids

Clinical and organizational factors associated with mortality during the peak of first COVID-19 wave: the global UNITE-COVID study

Purpose: To accommodate the unprecedented number of critically ill patients with pneumonia caused by coronavirus disease 2019 (COVID-19) expansion of the capacity of intensive care unit (ICU) to clinical areas not previously used for critical care was necessary. We describe the global burden of COVID-19 admissions and the clinical and organizational characteristics associated with outcomes in critically ill COVID-19 patients. Methods: Multicenter, international, point prevalence study, including adult patients with SARS-CoV-2 infection confirmed by polymerase chain reaction (PCR) and a diagnosis of COVID-19 admitted to ICU between February 15th and May 15th, 2020. Results: 4994 patients from 280 ICUs in 46 countries were included. Included ICUs increased their total capacity from 4931 to 7630 beds, deploying personnel from other areas. Overall, 1986 (39.8%) patients were admitted to surge capacity beds. Invasive ventilation at admission was present in 2325 (46.5%) patients and was required during ICU stay in 85.8% of patients. 60-day mortality was 33.9% (IQR across units: 20%–50%) and ICU mortality 32.7%. Older age, invasive mechanical ventilation, and acute kidney injury (AKI) were associated with increased mortality. These associations were also confirmed specifically in mechanically ventilated patients. Admission to surge capacity beds was not associated with mortality, even after controlling for other factors. Conclusions: ICUs responded to the increase in COVID-19 patients by increasing bed availability and staff, admitting up to 40% of patients in surge capacity beds. Although mortality in this population was high, admission to a surge capacity bed was not associated with increased mortality. Older age, invasive mechanical ventilation, and AKI were identified as the strongest predictors of mortality

Breast cancer management pathways during the COVID-19 pandemic: outcomes from the UK ‘Alert Level 4’ phase of the B-MaP-C study

Abstract: Background: The B-MaP-C study aimed to determine alterations to breast cancer (BC) management during the peak transmission period of the UK COVID-19 pandemic and the potential impact of these treatment decisions. Methods: This was a national cohort study of patients with early BC undergoing multidisciplinary team (MDT)-guided treatment recommendations during the pandemic, designated ‘standard’ or ‘COVID-altered’, in the preoperative, operative and post-operative setting. Findings: Of 3776 patients (from 64 UK units) in the study, 2246 (59%) had ‘COVID-altered’ management. ‘Bridging’ endocrine therapy was used (n = 951) where theatre capacity was reduced. There was increasing access to COVID-19 low-risk theatres during the study period (59%). In line with national guidance, immediate breast reconstruction was avoided (n = 299). Where adjuvant chemotherapy was omitted (n = 81), the median benefit was only 3% (IQR 2–9%) using ‘NHS Predict’. There was the rapid adoption of new evidence-based hypofractionated radiotherapy (n = 781, from 46 units). Only 14 patients (1%) tested positive for SARS-CoV-2 during their treatment journey. Conclusions: The majority of ‘COVID-altered’ management decisions were largely in line with pre-COVID evidence-based guidelines, implying that breast cancer survival outcomes are unlikely to be negatively impacted by the pandemic. However, in this study, the potential impact of delays to BC presentation or diagnosis remains unknown

Early mobilisation in critically ill COVID-19 patients: a subanalysis of the ESICM-initiated UNITE-COVID observational study

Background Early mobilisation (EM) is an intervention that may improve the outcome of critically ill patients. There is limited data on EM in COVID-19 patients and its use during the first pandemic wave. Methods This is a pre-planned subanalysis of the ESICM UNITE-COVID, an international multicenter observational study involving critically ill COVID-19 patients in the ICU between February 15th and May 15th, 2020. We analysed variables associated with the initiation of EM (within 72 h of ICU admission) and explored the impact of EM on mortality, ICU and hospital length of stay, as well as discharge location. Statistical analyses were done using (generalised) linear mixed-effect models and ANOVAs. Results Mobilisation data from 4190 patients from 280 ICUs in 45 countries were analysed. 1114 (26.6%) of these patients received mobilisation within 72 h after ICU admission; 3076 (73.4%) did not. In our analysis of factors associated with EM, mechanical ventilation at admission (OR 0.29; 95% CI 0.25, 0.35; p = 0.001), higher age (OR 0.99; 95% CI 0.98, 1.00; p ≤ 0.001), pre-existing asthma (OR 0.84; 95% CI 0.73, 0.98; p = 0.028), and pre-existing kidney disease (OR 0.84; 95% CI 0.71, 0.99; p = 0.036) were negatively associated with the initiation of EM. EM was associated with a higher chance of being discharged home (OR 1.31; 95% CI 1.08, 1.58; p = 0.007) but was not associated with length of stay in ICU (adj. difference 0.91 days; 95% CI − 0.47, 1.37, p = 0.34) and hospital (adj. difference 1.4 days; 95% CI − 0.62, 2.35, p = 0.24) or mortality (OR 0.88; 95% CI 0.7, 1.09, p = 0.24) when adjusted for covariates. Conclusions Our findings demonstrate that a quarter of COVID-19 patients received EM. There was no association found between EM in COVID-19 patients' ICU and hospital length of stay or mortality. However, EM in COVID-19 patients was associated with increased odds of being discharged home rather than to a care facility. Trial registration ClinicalTrials.gov: NCT04836065 (retrospectively registered April 8th 2021)

The Activation of Protein Kinase C and Protein Kinase D in Human Natural Killer Cells: The Effects of Tributyltin, Dibutyltin, and Tetrabromobisphenol A

Natural killer (NK) cells are lymphocytes that destroy (lyse) tumor cells, virally infected cells, and antibody coated cells. Although past studies have shown that inhibition of protein kinase C (PKC) blocks the lytic function of NK cells, the direct activation of PKC by exposure to lysis sensitive targets had not been examined. To verify that PKC and protein kinase D (PKD) activation occurs as a part of the lytic signaling pathway of NK cells, human NK cells were exposed to target tumor cells for 10 min, 30 min, and 1h. Exposure to K562 tumor cells for both 10 and 30 minutes caused an increase in the phosphorylation (activation) of both PKC and PKD. Previous studies have shown that the compounds tributyltin (TBT), dibutyltin (DBT), and tetrabromobisphenol A (TBBPA) decrease human NK cell lytic function and activate the mitogen activated protein kinase (MAPK) signaling pathway. The current study will examine the effects of DBT, TBT, and TBBPA exposures on the activation of PKC and PKD in human NK cells. NK cells were exposed to 300 - 25 nM TBT, 10 - 0.5 µM DBT, and 10 - 0.5 µM TBBPA for 10 min, 1h, and 6h. Ten minute exposures to 300 - 50 nM TBT activated PKC and to 300 - 100 nM TBT activated PKD; and significantly increased total PKD levels at 25 nM TBT. A 6h exposure to 300 - 25 nM TBT increased P-PKD levels but were not statistically significant. Ten minute exposure to 2.5 µM DBT and one hour exposure to 2.5 µM and 1µM DBT activated PKC. A 10 min exposure to 5 µM DBT activated PKD, and 0.5 µM DBT significantly decreased total PKD levels. The total PKD levels decreased significantly after 6h exposure to 10 and 5 µM TBBPA. P-PKC levels significantly decreased after a 6h exposure to 5 µM TBBPA. These data show for the first time that activation of PKD is part of the lytic signaling pathway of NK cells. Further, they demonstrate that exposure to the contaminants TBT and DBT activates the upstream activator of the lytic pathway, PKC as well as PKD

Retrospective evaluation of frozen section use for thyroid nodules with a prior fine needle aspiration diagnosis of Bethesda IIâVI: The Weill Cornell Medical College experience

Objective: To evaluate the Weill Cornell Medical College (WCMC)/New York Presbyterian Hospital (NYPH) experience with intraoperative frozen (IOF) section in the management of thyroid nodules with a fine needle aspiration (FNA) diagnosis of Bethesda IIâVI and to analyze the cost and pathology benefit it provides. Methods: The surgical and cytopathology files at WCMC/NYPH were searched within the time period of January 2008 to May 2013. A total of 435 thyroid specimens were identified for which both an FNA and subsequent IOF section was performed. The FNA was correlated with the locations of the resected nodule and the nodule frozen for intraoperative diagnosis. The results of the FNA were compared to the IOF section diagnosis and final diagnosis (FD). Results: Among 435 cases, the FNA diagnosis was Bethesda II: 149 cases, Bethesda III: 170 cases, Bethesda IV: 91 cases, Bethesda V: 19 cases, and Bethesda VI: 6 cases. There were a total of 83 carcinomas identified on FD, which included 69 papillary thyroid carcinomas (PTCs), 12 follicular carcinomas, and 2 poorly differentiated carcinomas. The preoperative FNA diagnosis for these carcinomas was as follows: Bethesda II, 11/149 (7.4%), Bethesda III, 24/170 (14%), Bethesda IV, 26/91 (29%), Bethesda V, 16/19 (84%), and Bethesda VI, 6/6 (100%). IOF section contributed to the diagnosis of malignancy in 16/429 (4%) cases: 1/149 (0.7%) Bethesda II, 5/170 (3%) Bethesda III, 2/91 (1.1%) Bethesda IV, and 8/19 (42%) Bethesda V. The diagnosis of malignancy was confirmed in the 6 Bethesda VI cases by IOF section. There were no false positives on IOF section. IOF had a sensitivity and specificity of 26% and 100%, respectively. Conclusion: The role of IOF section is limited in the evaluation of thyroid nodules. IOF section is most useful for nodules with an FNA diagnosis of Bethesda V lesions. The diagnosis of follicular variant of PTC remains difficult on frozen section. Keywords: Thyroid, Nodule, Frozen section, Fine-needle aspiration, Sensitivity, Specificit