bluuebunny/crossref_metadata_2025_split huggingface dataset:
Neutrophil‐Derived Microvesicles Enhance Pulmonary Vascular Inflammation via a Toll‐like Receptor 4 Signaling‐Dependent Mechanism
Authors: Tsiridou Diianeira Maria , Sachouli Eirini , Takata Masao , O'Dea Kieran P. 
Published: May 2022

BackgroundCirculating neutrophil‐derived microvesicles (NMVs) are markedly elevated during sepsis and therefore could have a role in the development of indirect acute lung injury (ALI). We recently found that NMV‐enriched CD11b+ MVs, immunoaffinity isolated from lipopolysaccharide (LPS)‐stimulated healthy volunteer blood, have potent pro‐inflammatory activity in a human peripheral blood mononuclear cell (PBMC) and lung microvascular endothelial cell (HLMEC) coculture model of pulmonary vascular inflammation (1). By contrast, immunoaffinity isolated platelet‐MVs (CD61+) produced negligible responses in these assays, suggesting specificity of the NMV‐enriched CD11b+ MV, activity. Here, we investigated the signaling mechanisms responsible for NMV‐mediated activation of monocytes and HLMECs in this model.MethodsHeparinized blood from healthy donors was treated with LPS (100 ng/ml, 3 h) and NMVs (CD66b+) were isolated by positive immunoaffinity selection. NMVs were then incubated in PBMC‐HLMEC cocultures for 4 h. NMV‐induced responses were determined by flow cytometric quantification of cell surface activation markers (ICAM‐1 for HLMECs, ICAM‐1 and tissue factor for monocytes) and pro‐inflammatory cytokine release by ELISA (TNFα, IL‐8, IL‐6, MCP‐1).ResultsNMV‐induced HLMEC activation required the presence of monocytes and was completely attenuated by anti‐TNFα neutralizing antibody (ICAM‐1: isotype control 302±103 vs anti‐TNFα 66±29; p<0.01). Pharmacological inhibition of mitogen‐activated protein kinases p38, MEK1‐MEK2 and phosphoinositide‐3 kinase ablated monocyte and HLMEC activation. Monocyte activation was prevented in the presence of the Toll‐like receptor 4 (TLR4) inhibitor TAK‐242 (e.g., TNFα release: vehicle 1108±257 vs TAK‐242 16±12; p<0.001) and partially inhibited by an anti‐TLR4 neutralizing antibody. However, the LPS inhibitor polymyxin‐B had no effect on these responses, indicating that LPS contamination was unlikely. Furthermore, NMV activity was found to be sensitive to proteinase K and heat‐treatment (55 °C, 15 min) (e.g., TNFα release: untreated 474±267 vs proteinase K 12±15; p<0.0001 vs heat‐treatment 200±84; p<0.01).ConclusionsWe have demonstrated that NMVs produced within LPS‐stimulated blood are potent mediators of lung microvascular endothelial cells via monocyte‐derived TNFα signaling. NMV‐induced monocyte activation involves TLR4 signaling and is protease sensitive, suggesting that MVs serve as vehicles of damage‐associated molecular pattern (DAMP) activity, mediated by surface proteins. Together these findings support a major role for NMVs as novel long‐range mediators for systemic propagation of inflammation, contributing to the pathogenesis of sepsis‐induced indirect acute lung injury.(1) Tsiridou et al. European Respiratory Journal. 2020;56:4468
A‐Kinase anchoring protein 2 (AKAP2) and Pendrin; cross‐talk between cell signaling and renal physiology
Authors: Belaid Houwayda , Wang Dongmei , Mendes Carraco Ana , Li Na , Van Bemmelen Miguel , Diviani Dario , Staub Olivier 
Published: May 2022

Kidney tubules play a pivotal role in the maintenance of body fluid homeostasis and pH regulation. Accordingly, kidney disorders are known to be strongly associated with hypertension and acid‐base imbalance. In the kidney, the intercalated cells of the nephron are the main site of acid‐base balance: type A are acid‐secreting whereas type B are base‐secreting. The chloride bicarbonate exchanger Pendrin is present in type B intercalated cells and participates actively in regulating blood pressure and NaCl balance. It is still not entirely deciphered how Pendrin activity is regulated; nevertheless, it was previously demonstrated that Pendrin is regulated by c‐AMP/Protein Kinase A (PKA) signaling pathway. Consequently, in this study, our hypothesis is that the A‐Kinase anchoring protein 2 (AKAP2), strongly expressed in the kidney, regulates in time and space the intracellular signal transduction.In order to investigate AKAP2 involvement in Pendrin regulation and trafficking to the apical plasma membrane, we generated an inducible and nephron‐specific Akap2Pax8/LC1 mice knockout model. By confocal microscopy, fluorescent immunostaining on kidney tissue sections showed that AKAP2 is present in the tubules and most importantly colocalizes with Pendrin at the apical plasma membrane of the intercalated cells. To show that Pendrin and AKAP2 are interacting, we used the Proximity Ligation Assay (PLA) and, positively, we could reveal the association between the two proteins. The latter finding was also confirmed in vitro taking advantage of the Opossum Kidney Cell line (OKP) stably expressing Pendrin and co‐transfected with AKAP2, we could co‐immunoprecipitate Pendrin and AKAP2 after crosslinking on live cells. Fluorescent immunostaining in Akap2Pax8/LC1 show that Pendrin tends to be shifted from the apical membrane (seen in control mice) to intracellular compartments.In conclusion, our data suggest that AKAP2 and Pendrin are interacting, and AKAP2 may play a key role in Pendrin trafficking to the apical plasma membrane, hence in regulating its activity.
Mycoplasma infection modifies attachment, proliferation, and the expression of immune checkpoint antigen CD276 (B7‐H3) on the Cal29 bladder cancer cell line
Authors: Aicher Wilhelm K. 
Published: May 2022

Introduction and aimBladder cancer is among the most prevalent malignancies in Western societies. Despite intensive research, progress towards better diagnosis or therapy was sparse in the last decades. However, many studies documented elevated expression of immune checkpoint antigens including PD‐1, PD‐L1, CTLA‐4, or CD276 (alias B7‐H3) on bladder tumor cells. Immune checkpoint antigens are capable to suppress immune responses against the tumor. But novel therapies targeting immune checkpoint antigens are promising. Some of these agents are already part of clinical studies. At the same time mycoplasma infections were reported in about 18% of sexually experienced adults. We therefore investigated if mycoplasma contaminated urothelial carcinoma cells Cal29 differed in attachment, appearance, proliferation, and expression of immune checkpoint antigen CD276 from mycoplasma‐free Cal29.Materials and MethodsMycoplasma infected Cal29 cells (mcCal29) and clean Cal29 cells (Cal29, from ATCC) were expanded and infection vs. sterility was monitored by PCR throughout the experiment. Cells were seeded and attachment, appearance and size of the cells were observed by microscopy. Proliferation was enumerated in consecutive passages using a hematocytometer and trypan blue dye exclusion. Expression of CD276 was studies on the mRNA level by quantitative PCR of cDNA (RT‐qPCR). Analyses of GAPDH and PPIAγ served as controls. CD276 protein levels were explored by immunoblot, and ß‐actin served a control. CD276 expression on cell surfaces was explored by flow cytometry and is documented as mean fluorescence intensity (MFI). Unstained cells and beads served as controls.ResultsMycoplasma‐contaminated Cal29 proliferated significantly faster than clean Cal29 (n=5, 132%, p<7,8E‐09). Upon seeding in flasks, Cal29 adhered faster then mcCal29, attached more firm to the flasks as microscopy showed up to double diameters in comparison to mcCal29. Expression of CD276 transcripts and total proteins did not differ, but on cell surfaces mcCal29 (MFI 4334) expressed less than half of CD276 in comparison to Cal29 (MFI 9729).ConclusionsUrinary infection by mycoplasma may indirectly contribute to tumor spready by accelerating proliferation of bladder cancer cells and by reducing cell‐cell and/or cell‐matrix interactions as documented by attachment and cell growth studies. However, mcCal29 presented less CD276 on tumor cell surfaces. Thus, mycoplasma infection may facilitate immune responses to bladder cancer cells by reducing the expression of CD276. But these preliminary results must be conformed or challenged by additional studies with other urothelial cells.
Jadomycin B Acts Synergistically with NSAIDs to Produce Cytotoxic Effects in Human Breast Cancer Cells in vitro
Authors: McKeown Brendan T. , Goralski Kerry B. 
Published: May 2022

BackgroundBreast cancer is the most prevalent cancer in North American women. Jadomycin B is a novel compound that is effective in killing triple negative human breast cancer cells that have become resistant to anthracycline and taxane chemotherapies. The mechanism through which jadomycin B exerts its cytotoxic effect remains to be fully characterized. A quantitative PCR (qPCR) array of cancer drug targets identified cyclooxygenase‐2 (COX‐2) as a gene of interest in jadomycin B resistant breast cancer cells.ObjectiveTo explore the effects of COX‐2 signalling on jadomycin B cytotoxicity in MDA‐MB‐231 cells as a model of invasive triple‐negative, human breast cancer requiring more effective treatments.Methods and ResultsControl MDA‐MB‐231 (231‐CON) cells exposed to increasing concentrations of jadomycin B (0‐2.2 µM) over 7 months developed a 3‐fold resistance to jadomycin B as determined using methyl‐tetrazolium (MTT) cell viability assays. By qPCR, there was a significant increase in COX‐2 (40 fold, p < 0.05) but not cyclooxygenase‐1 (COX‐1) (0.17 fold, p < 0.05), mRNA expression in jadomycin resistant (231‐JB) versus 231‐CON cells. A similar increase in COX‐2 was not observed in mitoxantrone resistant MDA‐MB‐231 cells (231‐MITX). We therefore hypothesized that inhibition of COX‐2 signalling would increase the susceptibility of breast cancer cells to jadomycin B. The 231‐CON and 231‐JB cells were treated with jadomycin B with or without the COX‐2 selective inhibitor celecoxib (CXB), or COX‐1/2 inhibitors ibuprofen (IBU) or Naproxen (NAP). The SynergyFinder 2.0 web application was used to analyze the cytotoxicity (MTT assays) of these combinations and calculate synergy scores using the Bliss model. IBU and NAP were found to act synergistically with jadomycin B in both control and jadomycin resistant cells, while CXB acted additively (Table 1).ConclusionsJadomycin B acts additively or synergistically with inhibitors of COX‐2 in vitro, resulting in increased cytotoxic effect. These results justify further investigation into the COX‐2 signaling pathway as a means to elucidate the mechanism of action of jadomycin B. Additionally, our research has the potential to contribute to an improved understanding of breast cancer relevant COX‐2 signalling, facilitating the use of COX‐2 inhibitors as chemosensitizers.
Laminar Shear Stress Enhances Cytosolic PINK1 Expression: Increased Mitophagic Sensitivity Toward Mitochondrial Dysfunction
Authors: Hong Soon‐Gook , Shin Junchul , Saredy Jason , Choi Soo Young , Wang Hong , Park Joon‐Young 
Published: May 2022

PURPOSEPhosphatase and tensin homolog (PTEN)‐induced putative kinase 1 (PINK1) is an essential molecule in mitophagy process and known to act as a cytoprotective protein in mammalian cells. Mutation or deficiency of PINK1 has been closely related to several disease conditions. The purpose of this study was to determine PINK1 expression levels and subcellular localization under exercise‐mimic laminar shear stress (LSS) condition in human aortic endothelial cells (HAECs) or in exercising mice, and its implication on endothelial homeostasis and cardiovascular disease (CVD) prevention.METHODS & RESULTSFirst, we measured full‐length PINK1 (FL‐PINK1) mRNA and protein expression levels in HAECs under unidirectional laminar shear stress (LSS) at 15 dyne/cm2 for 48h. LSS significantly elevated both FL‐PINK1 mRNA and protein expressions in HAECs compared to static control. Mitochondrial fractionation assays showed a decrease in FL‐PINK1 accumulation in the mitochondria with a compensatory increase in the cytosolic FL‐PINK1 level under LSS. Confocal microscopic analysis confirmed these subcellular localization patterns suggesting downregulation of mitophagy induction. Also, mitophagy flux was decreased under LSS, determined by a mtKeima probe. Mitochondrial morphometric analysis and mitochondrial membrane potential (JC‐1) showed mitochondrial elongation and increased mitochondrial membrane potential, respectively, suggesting that an elevation of cytosolic PINK1 is not related to an immediate induction of mitophagy. Based on this observation, we further hypothesized that the elevation of the cytosolic pool of FL‐PINK1 would increase the sensitivity to mitophagic signals in pathological conditions. Preconditioned HAECs with LSS showed lower mtDNA lesions under angiotensin II stimulation (100 nM, 6h). Moreover, LSS‐preconditioned ECs showed rapid Parkin recruitment and mitophagy induction in response to mitochondrial toxin (i.e. CCCP) treatment compared to the control. We measured PINK1 expression at ECs of the thoracic aorta in exercised mice, a physiological LSS‐enhanced model, which was significantly elevated compared to sedentary animals. In addition, exercise‐preconditioned mice were more protective to angiotensin II‐induced mtDNA lesion formation in the mouse abdominal aorta than sedentary mice, suggesting a potential protective mechanism of exercise in a PINK1‐dependent manner.CONCLUSIONLSS increases a cytosolic pool of FL‐PINK1, which may elevate the mitophagic sensitivity toward dysfunctional mitochondria or activate other cytoprotective mechanisms in endothelial cells. Our data suggest that exercise may support mitochondrial homeostasis in vascular endothelial cells by enhancing PINK1‐dependent cell protection mechanisms.
Acute Respiratory Muscle Fatigue in Patients with Obesity Following Peak Aerobic Exercise
Authors: Severin Richard S. , Alshammari Saleh , Sherman Sara R. , Phillips Shane A. 
Published: May 2022

IntroductionRespiratory muscle (RM) performance has been theorized to contribute to exercise‐induced dyspnea, exercise intolerance, and low exercise capacity in patients with obesity. However, the impact of acute impairments in RM performance on exercise intolerance and exercise capacity in patients with obesity remains uncertain. We hypothesized, that following peak aerobic exercise, impairments in RM performance would be observed in patients with obesity.MethodsWe prospectively recruited 7 participants (recruitment is ongoing; 1 male, 6 female, Age: 42 ± 9 yrs; BMI: 47.7 ± 5.9 kg/m2; Waist Circumference: 126 ± 12.04 cm; Body Fat [%]: 50.91 ± 3.6). Diaphragm Thickness at expiration and Thickening Ratio (thickness at end‐inspiration – thickness at end‐expiration)/thickness at end‐expiration) were assessed via B‐mode ultrasound at the right 8th intercostal space while patients were in a semi‐recumbent position elevated 30 degrees from supine. Participants performed a cardiopulmonary exercise test (CPET) using a ramp treadmill protocol to peak exercise capacity. Measurements of RM performance were measured at baseline and repeated at 0‐minutes and 5‐minutes following CPET; which included Maximal Inspiratory Pressure (MIP), Sustained Maximal Inspiratory Pressure (SMIP), Inspiratory Duration (ID), Slope of the SMIP Plot (SMIP Slope), and Maximal Expiratory Pressure (MEP). Two‐Sided Paired T‐Tests were used to compare RM performance data before and after testing; statistical significance was set at p<0.05.ResultsMean CPET Results: VO2peak: 19.37 ± 2.4 mL/kg/min; RER: 1.01 ± 0.11; Age Predicted Heart Rate Max [%] 87.57 ± 0.14; Ventilatory Reserve [%] 25.25 ± 18.32; VE/VCO2 Slope: 28.84 ± 4.50. Diaphragm Thickness: 2.8 ± 0.24 mm and Thickening Ratio: [%]: 75.47 ± 25.97. When comparing Baseline to 0‐minues following CPET significant reductions in MIP, SMIP, and MEP were observed: MIP (94.57 ± 25.01 cmH2O vs. 72.29 ± 22.09 cmH2O p = 0.02); SMIP (524.14 ± 198.85 PTU vs. 418.43 PTU p = 0.04); MEP (73.71 ± 17.10 cmH2O vs. 62.29 ± 12.83 cmH2O p= 0.02). A reduction in ID (10.6 ± 2.30 vs 8.99 sec p= 0.06) and an increase in SMIP Slope (9.2 ± 3.39 vs. 9.74 ± 3.30 p= 0.74) were observed at 0‐minutes following CPET compared to baseline but were not significant. At 5‐minutes following CPET, significant reductions in MIP (67.3 cmH2O ± 12.6 p= 0.01) and MEP (62.57 ± 14.47 cm H2O p= 0.01) were observed but not for other measures of RM performance.ConclusionOur results suggest that patients with obesity demonstrate significant reductions in RM performance following peak aerobic exercise which may persist even after recovery. Interestingly, at baseline, the patients with obesity in this study demonstrated relatively normal RM performance, diaphragm thickness and thickening ratio. Acute fatigue of the respiratory muscles may contribute to the exercise induced dyspnea, exercise intolerance and low exercise capacity observed in patients with obesity.
Correlation Between Liver Weight and Dimensions to Patient Demographics
Authors: Alimohammadi Majid 
Published: May 2022

BackgroundTreatment of hepatocellular carcinoma (HCC) often involves the use of targeted radiation therapy, chemotherapeutic agents, and percutaneous ethanol injections, but issues of underdosing and overdosing still prevail and dosimetric calculations are often suboptimal.Methods33 cadavers donated to the University of British Columbia (UBC) Body Donation Program were examined, and data was collected on liver weight and dimensions. The individual’s demographics (height, weight) were obtained from the department’s database, and the liver demographics were correlated with the individuals’ demographics.ResultsData was analyzed for correlations between body weight and liver weight, person’s height and the height of the liver along the right surface, body weight and the liver’s length along the superior surface, and body weight and the distance the liver extended left of the midline. A strong positive correlation was identified between body weight and liver weight, with r=0.7475 and p < 0.00001. All other results either concluded weak correlations, or were insignificant.DiscussionThe strong correlation identified between body weight and liver weight can be used as a potential additional tool during initial clinical examinations to aid dosimetric calculations of radiation and chemotherapy treatments.
N‐Glycosylation Inhibitor Interferes with WNT Signaling and Inhibits Angiogenesis
Authors: Banerjee Dipak K. , Lebron Arelis S. , Collazo Irizarry Diego K. , Baksi Krishna 
Published: May 2022

Neovascularization, a normal physiological process orchestrates through vasculogenesis from mesoderm cell precursors. In healthy adults, the vasculature is quiescent but pathological angiogenesis is evident during wound healing and in tumor growth. Thus, angiogenesis becomes a fundamental step for tumors transitioning from a benign state to malignancy where endothelial cells proliferate 45 times faster. Our laboratory has recently shown that asparagine‐linked (N‐linked) protein glycosylation is essential for enhanced capillary endothelial cell proliferation in situ. Inhibiting N‐glycosylation with Tunicamycin causes G1 arrest and induces cell death by apoptosis due to “ER stress” and is not reversible by VEGF. Tunicamycin also inhibits angiogenesis in vivo in Matrigel™ implants in nude mice as well as in a pre‐clinical mouse model of double negative (ER‐/PR‐/HER2+) breast tumor.Wnt pathway is involved in cellular differentiation to organogenesis, and supports angiogenesis. In “canonical” Wnt/β‐catenin signaling pathway β‐catenin translocates to nucleus and activates Wnt/β‐catenin target genes, including vascular endothelial growth factor A (VEGF‐A). Irrespective of the Wnt ligand and Frizzled being glycoproteins, the cross‐talk between protein N‐glycosylation and the Wntpathway has not been addressed. This led us to hypothesize that decreased expression of Wntprotein(s) in Tunicamycin treated endothelial cell would decrease cellular proliferation. We have used capillary endothelial cells as a model and observed down regulated expression of Wnt proteins in Tunicamycin treated cells. Our results support Tunicamycin‐induced “ER stress” interferes with Wnt signaling and inhibits angiogenesis. We, therefore, conclude molecular understanding of the process would provide a novel target to inhibit angiogenesis and breast tumor progression.
Soft silicone‐based neural interface to modulate bladder function: chronic studies in awake behaving cats
Authors: Kumar Ritesh , Gopinath Chaitanya , Simpson Tyler W. , Weir David M. , Thiessen Alexander , McDonnall Danny , Gaunt Robert A. 
Published: May 2022

Direct bladder wall stimulation has been attempted for decades to restore bladder function in people with spinal cord injury and other voiding dysfunctions. However, these efforts were limited by co‐activation of the urethra, legs and other pelvic organs at stimulus intensities that evoked bladder contractions. Neural interfaces for the detrusor muscle itself face several challenges due to its structure and the volume changes it undergoes in normal function. We designed a stretchable silicone net that can be placed around the bladder body to anchor a soft electrode array that interfaces directly with the base of bladder to generate bladder contractions.We created implantable versions of the electrode nets and tested them in chronic experiments. We implanted 5 healthy cats (4 females, 1 male) and tested them with and without anesthesia for 2‐3 months. A pressure catheter was implanted into the bladder to control volume and measure pressure.Bladder wall stimulation through various electrode configurations (monopolar, bipolar), temporal patterns (single electrode, sequential stimulation of multiple electrodes) and stimulus intensities were able to generate complete bladder emptying. In behaving cats, bladder wall stimulation at many different stimulus intensities elicited efficient voiding at physiological bladder pressures (med: 41, IQR: 27‐60 cmH2O). However, there was little relationship (r<0.28) between stimulation amplitude and bladder pressure itself. Rather, stimulation amplitude had an effect on the time between stimulation onset and bladder contraction onset. At low stimulation amplitudes, there was a longer time between stimulation onset and voiding (med: 35s, IQR: 13.5‐68.2s) than occurred at higher amplitudes, which evoked more rapid voiding (med: 23s, IQR: 10‐45s). This contrasts with anesthetized tests, where stimulation at 3 ± 2 mA evoked increases in bladder pressure up to 35 cmH2O. These results suggest that stimulation of the bladder wall recruits sensory pathways leading to an urge to void, but that can be suppressed by the animals. For most of the duration of the implants, stimulation evoked functional voiding with consistent electrical stimulation thresholds. However, after several months there was a failure of an implanted bond that limited the overall duration of the implants.Chronic experiments demonstrate that these electrode nets can be used as a neural interface to generate or initiate comfortable, complete bladder emptying in awake behaving cats. Future efforts include understanding the effects of evoked sensations on voiding and improving the integrity of the bonding between the flexible conductors and the lead wire.