Thromboxane A2 and TP receptors: A trail of research, well traveled

This article was written is response to a request from the editor-in-chief as a result of my receiving the Governor’s Award in Scientific Awareness. My goal is to provide a glimpse into a 40 plus year research career that was predominately focused on the role of thromboxane A2 and its receptor (TP) in physiologic and pathophysiologic processes in the cardiovascular system. It also enumerates the lessons learned along the way that may lead to a productive research career. Those lessons were; Perseverance, Think out of the box, Challenge the dogma, Take risks, Failure is not a bad thing, Focus, Select a good mentor(s), Choose your collaborators and colleagues wisely (team science), When you start an experiment-finish it (analyze all the data), Take time for your family

Thromboxane and Stable Prostaglandin Endoperoxide Analogs Stimulate Water Permeability in the Toad Urinary Bladder

The effects of thromboxane B2 and the stable prostaglandin endoperoxide analogs (15Z)-hydroxy - 9α - 1 lα - (epoxymethano)prosta - 5Z,13E – dienoic acid (U44069) and (15Z)-hydroxy-1 lα,9α-(epoxymethano) prosta-5Z,13E-dienoic acid (U46619) were tested on water flow across the toad urinary bladder. In the presence of indomethacin or meclofenamic acid, inhibitors of prostaglandin and thromboxane A2 synthesis, thromboxane B2 stimulated water flow in a dose-dependent manner. U44069 (1 µM) stimulated water flow from 3.6±0.8 to 12.4±1.2 mg/min per 10 cm2 hemibladder surface area, while U46619 (1 µM) stimulated water flow from 2.8± 1.0 to 21.8±2.0 mg/min per 10 cm2. The prostaglandin endoperoxide/thromboxane A2 antagonist trans-13-azaprostanoic acid, an inhibitor of vasopressin-stimulated water flow, inhibited thromboxane B2- and U46619-stimulated water flow in a dose-dependent manner. The inactive cis-13-azaprostanoic acid did not inhibit vasopressin-stimulated water flow in untreated hemibladders and had no effect on U46619-stimulated water flow in indomethacin or meclofenamic acid pretreated hemibladders. U46619 (1 µM) enhanced vasopressin-stimulated water flow in indomethacin pretreated hemibladders, producing a significant parallel shift (P \u3c 0.001) in the dose-response relationship to submaximal concentrations of vasopressin (0.1-0.6 mU/ml), while not affecting water flow stimulated by supramaximal concentrations of vasopressin (10 mU/ml). trans-13-Azaprostanoic acid abolished the potentiating effects of U46619 on vasopressin-stimulated water flow. These results show that thromboxane A2-like compounds stimulate water flow in the toad urinary bladder

Elevated Thromboxane Levels in the Rat during Endotoxic Shock: Protective Effects of Imidazole, 13-azaprostanoic Acid, or Essential Fatty Acid Deficiency

The potential deleterious role of the proaggregatory vasoconstrictor, thromboxane A2, in endotoxic shock was investigated in rats. Plasma thromboxane A2 was determined by radioimmunoassay of its stable metabolite thromboxane B2. After intravenous administration of Salmonella enteritidis endotoxin (20 mg/kg), plasma thromboxane B2 levels increased from nondetectable levels (\u3c375 pg/ml) in normal control rats to 2,054±524 pg/ml (n = 8), within 30 min to 2,071+429 at 60 min, and decreased to 1,119+319 pg/ml, at 120 min. Plasma levels of prostaglandin E also increased from 146±33 pg/ml in normal controls (n = 5) to 2,161+606 pg/ml 30 min after endotoxin (n = 5). In contrast to shocked controls, rats pretreated with imidazole, a thromboxane synthetase inhibitor, or essential fatty acid-deficient rats, which are deficient in arachidonate and its metabolites, did not exhibit significant elevations in plasma levels of thromboxane B2. Imidazole did not however inhibit endotoxin-induced elevations in plasma prostaglandin E. Essential fatty acid deficiency significantly reduced mortality to lethal endotoxic shock. This refractoriness could be duplicated in normal rats pretreated with the fatty acid cyclo-oxygenase inhibitor, indomethacin (10 mg/kg), intravenously 30 min before endotoxin injection. Imidazole (30 mg/kg) administered intraperitoneally 1 h before or intravenously 30 min before endotoxin, also significantly (P \u3c 0.01) reduced mortality from lethal endotoxin shock to 40% compared to a control mortality of 95% at 24 h. Likewise pretreatment with 13-azaprostanoic acid (30 mg/kg), a thromboxane antagonist, reduced mortality from endotoxic shock at 24 h from 100% in control rats to only 50% (P \u3c 0.01). The results suggest that endotoxin induces increased synthesis of thromboxane A2 that may contribute to the pathogenesis of endotoxic shock

交感神経β受容体遮断薬propranololの降圧機序に関する研究 : 血管拡張性prostaglandinの役割を中心に

β-adrenergic receptor antagonists are widely used to treat the patients with hypertension. The mechanism of the antihypertensive effect of these drugs was thought to be associated with 1) negative cardiac inotropic and chronotropic action, 2) suppression of renin secretion, and 3) effects on the central nervous system. However, the precise mechanism is still unknown. Long term therapy with β-adrenergic receptor antagonists is also associated with a reduction in the peripheral vascular resistance. Indomethacin and other non-steroidal antiinflammatory drugs have been shown to blunt the antihypertensive effect of propranolol and other β-adrenergic receptor antagonists in patients and laboratory animals. This study was, therefore, designed to determine the effects of DL-, D- or L-propranolol on the synthesis of the vasodilatory prostaglandin, PGI2 and PGE2 in the aorta, renal medulla, and the renal cortex of spontaneously hypertensive rats (SHR). DL-Propranolol and L-propranolol significantly lowered blood pressures from 148±9/113±5mmHg to 112±3/80±3 and from 133±4/100±2mmHg to 121±3/81±3 mmHg, respectively. Comparable treatment of SHR with D-propranolol revealed no lowering effect. Basal immunoreactive 6-keto PGF1α and PGE2 production by isolated thoracic aorta, renal medulla, and renal cortex was measured in the samples with an incubation time of 60 to 90 minutes and showed no difference in the vehicle compared to the DL-propranolol treated rats. Norepinephrine (1 μM)-stimulated synthesis of 6-keto PGF1α and PGE2 in the aorta was significantly enhanced in the DL-propranolol treated group compared to the vehicle treated group. Aortic 6-keto PGF1α, and PGE2 synthesis stimulated by norepinephrine in the L-propranolol treated rats was also significantly higher than that observed in the vehicle and D-propranolol treated groups. DL-propranolol treatment did not alter norepinephrine-stimulated renal cortical or medullary 6-keto PGF1α and PGE2 production. Bradykinin- and angiotensin II- augmented aortic 6-keto PGF1α along with PGE2 production in the chronic treatment with L-propranolol was significantly higher than that in the D-propranolol treated group. However, arachidonic acid-stimulated aortic 6-keto PGF1α and PGE2 production in the chronic treatment with DL-propranolol group was not different from that of the vehicle treated group. Acute in vitro exposure of thoracic aortas from SHR to D-, or L-propranolol (100ng/ml) did not ?augment aortic 6-keto PGF1α and PGE 2 synthesis compared to the vehicle. These data suggest that 1) the antihypertensive effect of propranolol is stereoselective, 2) norepinephrine-, bradykinin-, and angiotensin II-stimulated aortic vasodilatory prostaglandin synthesis are stereoselective, 3) the antihypertensive effect of propranolol may be related to the enhanced synthesis of vasodilatory prostaglandin, PGI2 or PGE2, in the vascular tissue, and 4) chronic DL-propranolol treatment may modulate arachidonic acid metabolism by changing either phospholipase A2 activity or the membrane phospholipids metabolism

Circulating extracellular vesicles are associated with the clinical outcomes of sepsis

IntroductionSepsis is associated with endothelial cell (EC) dysfunction, increased vascular permeability and organ injury, which may lead to mortality, acute respiratory distress syndrome (ARDS) and acute renal failure (ARF). There are no reliable biomarkers to predict these sepsis complications at present. Recent evidence suggests that circulating extracellular vesicles (EVs) and their content caspase-1 and miR-126 may play a critical role in modulating vascular injury in sepsis; however, the association between circulating EVs and sepsis outcomes remains largely unknown.MethodsWe obtained plasma samples from septic patients (n=96) within 24 hours of hospital admission and from healthy controls (n=45). Total, monocyte- or EC-derived EVs were isolated from the plasma samples. Transendothelial electrical resistance (TEER) was used as an indicator of EC dysfunction. Caspase-1 activity in EVs was detected and their association with sepsis outcomes including mortality, ARDS and ARF was analyzed. In another set of experiments, total EVs were isolated from plasma samples of 12 septic patients and 12 non-septic critical illness controls on days 1, and 3 after hospital admission. RNAs were isolated from these EVs and Next-generation sequencing was performed. The association between miR-126 levels and sepsis outcomes such as mortality, ARDS and ARF was analyzed.ResultsSeptic patients with circulating EVs that induced EC injury (lower transendothelial electrical resistance) were more likely to experience ARDS (p<0.05). Higher caspase-1 activity in total EVs, monocyte- or EC-derived EVs was significantly associated with the development of ARDS (p<0.05). MiR-126-3p levels in EC EVs were significantly decreased in ARDS patients compared with healthy controls (p<0.05). Moreover, a decline in miR-126-5p levels from day 1 to day 3 was associated with increased mortality, ARDS and ARF; while decline in miR-126-3p levels from day 1 to day 3 was associated with ARDS development.ConclusionsEnhanced caspase-1 activity and declining miR-126 levels in circulating EVs are associated with sepsis-related organ failure and mortality. Extracellular vesicular contents may serve as novel prognostic biomarkers and/or targets for future therapeutic approaches in sepsis

Organization of sorting and surgery of wounds with soft tissue defects during the joint force surgery

Introduction. The experience of providing medical care during the Anti-terrorist operation in eastern Ukraine showed that in the structure of modern combat surgical trauma gunshot wounds with soft tissue defects are between 64.9-68.2%, of which 36.4-37.5% are small and medium, 28.5-30.7% are large and very large defects.Aim: To improve the results of providing surgical care to the wounded with soft tissue defects by introducing a variety of surgical tactics of wound closure to the medical care levels.Material and Methods. The total array of the study was 2537 wounded with shrapnel, bullet and mine injuries from April 2014 to September 2018. The determination of surgical tactics for closing soft tissue defects was performed at the basis of metric classification taking into account the area, volume and anatomical areas of the lesion.Results. The combination of metric characteristics of wound defects by area, volume with localization of wounds in a single classification allowed the offer of a comprehensive approach to sorting the wounded at the level of medical care and to determine further surgical tactics to close soft tissue defects. In accordance with the sorting and evacuation purposes, the wounded with gunshot wounds to the foot and hand (third zone of injury) were treated in specialised centres to the fourth level of medical care. In the case of medium and large wounds of the thigh, leg, shoulder and forearm, medical care was provided at the second and third levels. And in the case of large and very large wounds of the specified localisation was provided in specialised clinics of the fourth level.Conclusions. The introduction of differentiated surgical tactics in the wounded with soft tissue defects at the levels of medical care has improved functional results: increase the proportion of good from 46.9% to 53.7%, reduce the relative number of unsatisfactory from 18.8% to 11, 6%

Мікробіота породних відвалів вугільних шахт Червоноградського гірничопромислового району за внесення золи

The aim of this work was to determine the impact of addition of coal ash from Dobrotvir TPP to waste heaps gangue (Chervonograd Mining Region) on the number of different groups of microorganisms. 20 samples from three waste heaps, from the black and red gangue, under the mosses and from bare substrate and also from terrace, top and base of each waste heap, were selected. Waste heaps gangues with coal ash from Dobrotir TPP were mixed in vitro and left for 10 days. We used proportion of coal ash to gangue as 1 to 5. Microorganisms were grown in Petri dishes containing 20–30 ml agar medium and in 22 ml tubes at temperature of 28 °C. Microscopic fungi were revealed on Mash-agar; oligonitrophilic bacteria – on Ashby medium; actinomycetes – on Chapek’s medium; cellulose decomposing aerobic bacteria – on Hetchenson medium; colorless sulfur oxidizing bacteria: neutrophilic – on Beyerinck medium, acidophilic – on Silverman and Lundgren 9К medium. The acidity value of waste heaps gangue samples was determined by рН meter рН-150М. We observed that samples collected under the mosses had lower acidity compared to samples from the bare substrate. We also revealed lower acidity of the overburn red gangue than the acidity of freshly deposited black gangue. To sum up, application of coal ash resulted in lowering of acidity value among all samples under study. Coal ash addition led to increase in number of microscopic fungi cells compared to the appropriate control samples. The highest quantity of microscopic fungi (16.2 ± 0.79) х 105 CFU/g of gangue) was revealed in sample from red rock of the main waste heap of Central Enrichment Plant (CEP). At the same time, we observed the highest cell number in the control sample under the mosses of “Nadija” coal pit waste heap, (6.1 ± 0.3) х 105 CFU/g of gangue. After coal ash addition, most samples featured 2–3 times higher quantities of colorless sulfur-oxidizing neutrophilic bacteria cells. The highest cell number of these microorganisms was observed in sample under the mosses of “Vizejska” coal pit waste heap. The same dominance of oligonitrophilic bacteria cell number in experimental samples over control samples was indicated. The highest cell quantity was recorded for the sample under the moss from terrace of waste heap of “Nadija” coal pit (25.6 ± 1.3) х 104 CFU/g of gangue and in sample from red rock of CEP main waste heap (21.1 ± 1.1) х 104 CFU/g of gangue, being 43.3 and 31.7 times higher than the appropriate controls. Meanwhile, the number of colorless sulfur-oxidizing acidophilic bacteria in control samples was higher than that after coal ash addition, particularly, on the bare substrate samples from the base and terrace of waste heap of “Nadija” coal pit. Higher cell number in control samples without coal ash was typical for actinomycetes with the greatest difference (8.6 times) before and after coal ash addition in a sample from red rock of CEP main waste heap. We detected 2 times lower number of cellulose-decomposing aerobic bacteria in the majority of experimental samples compared to appropriate control samples under study. In that way, we noticed that addition of coal ash from Dobrotvir TPP to waste heaps gangue (in proportion of coal ash to gangue as 1 to 5) caused reduction of substrate acidity value. Under these conditions the number of colorless sulfur-oxidizing neutrophilic bacteria, oligonitrophilic bacteria and microscopic fungi cells increased. But on the other hand, coal ash addition resulted in lowering of the number of colorless sulfur-oxidizing acidophilic bacteria, actinomycetes, and cellulose-decomposing aerobic bacteria cells. Досліджено мікробіоту породного відвалу Центральної збагачувальної фабрики, відвалів вугільних шахт «Візейська» та «Надія» Червоноградського гірничопромислового району. Показано вплив внесення золи із Добротвірської ТЕС до породи відвалів на кислотність субстратів та чисельність еколого-трофічних груп мікроорганізмів у лабораторних умовах. Внесення золи у пропорціях зола : порода 1 : 5 сприяє підлужненню субстратів відвалів, змінюючи кислотність на 1,25–2,59 одиниці порівняно з вихідними субстратами. Внесення золи до порід відвалів зумовило підвищення чисельності клітин мікроскопічних грибів порівняно з контролями з найвищою кількістю у пробах червоної породи основного відвалу Центральної збагачувальної фабрики (ЦЗФ). Без внесення золи найвищу чисельність виявили у пробах, узятих під мохом тераси відвалу шахти «Надія». У більшості проб після внесення золи чисельність клітин безбарвних сіркоокиснювальних нейтрофільних бактерій була удвічі-втричі вищою, ніж у контролях із найвищою кількістю клітин у дослідному зразку під мохом тераси відвалу шахти «Візейська». Для олігонітрофільних бактерій відмітили їх домінування у дослідних пробах із внесенням золи. Чисельність клітин безбарвних сіркоокиснювальних ацидофільних бактерій у контрольних пробах була вищою, ніж у пробах після внесення золи, зокрема в оголених субстратах проб із підніжжя та тераси відвалу шахти «Надія». Для актиноміцетів також виявили помітно вищу кількість у контрольних пробах без внесення золи з найістотнішою різницею у 8,6 раза у пробах червоної породи основного відвалу ЦЗФ. У більшості досліджених зразків внесення золи зумовило зниження кількості клітин деструкторів целюлози удвічі порівняно з контролями. Досліджено мікробіоту породного відвалу Центральної збагачувальної фабрики, відвалів вугільних шахт «Візейська» та «Надія» Червоноградського гірничопромислового району. Показано вплив внесення золи із Добротвірської ТЕС до породи відвалів на кислотність субстратів та чисельність еколого-трофічних груп мікроорганізмів у лабораторних умовах. Внесення золи у пропорціях зола : порода 1 : 5 сприяє підлужненню субстратів відвалів, змінюючи кислотність на 1,25–2,59 одиниці порівняно з вихідними субстратами. Внесення золи до порід відвалів зумовило підвищення чисельності клітин мікроскопічних грибів порівняно з контролями з найвищою кількістю у пробах червоної породи основного відвалу Центральної збагачувальної фабрики (ЦЗФ). Без внесення золи найвищу чисельність виявили у пробах, узятих під мохом тераси відвалу шахти «Надія». У більшості проб після внесення золи чисельність клітин безбарвних сіркоокиснювальних нейтрофільних бактерій була удвічі-втричі вищою, ніж у контролях із найвищою кількістю клітин у дослідному зразку під мохом тераси відвалу шахти «Візейська». Для олігонітрофільних бактерій відмітили їх домінування у дослідних пробах із внесенням золи. Чисельність клітин безбарвних сіркоокиснювальних ацидофільних бактерій у контрольних пробах була вищою, ніж у пробах після внесення золи, зокрема в оголених субстратах проб із підніжжя та тераси відвалу шахти «Надія». Для актиноміцетів також виявили помітно вищу кількість у контрольних пробах без внесення золи з найістотнішою різницею у 8,6 раза у пробах червоної породи основного відвалу ЦЗФ. У більшості досліджених зразків внесення золи зумовило зниження кількості клітин деструкторів целюлози удвічі порівняно з контролями.

Alterations in macrophage G proteins are associated with endotoxin tolerance

AbstractPrevious studies have suggested that endotoxin tolerance induces macrophage desensitization to endotoxin through altered guanine nucleotide regulatory (G) protein function. In the present study the binding characteristics of the nonhydrolyzable GTP analogue GTPγ[35S] to macrophage membranes from endotoxin tolerant and control rats were determined. Membranes were prepared from peritoneal macrophages harvested from rats 72 h after two sequential daily doses of vehicle or Salmonella enteritidis endotoxin (100 μg/kg on day 1 and 500 μg/kg on day 2). GTPγ[35S] bound to a single class of sites that were saturable and displaceable in control and endotoxin tolerant macrophage membranes. The maximum specific binding of GTPγ[35S] was significantly (P < 0.01) decreased in membranes from tolerant rats compared to control (Bmax = 39 ± 7 pmol/mg protein in control vs. 11 ± 2 pmol/mg protein in endotoxin tolerant; n = 5). There were no significant differences in the Kd values. To determine whether the reduced GTPγS binding was due to decreases in G proteins, macrophage membrane G protein content was determined by western blotting with specific antisera to Gi1,2 α, Gi3α, Gs α, and the β subunit of G. Scanning densitometric analysis demonstrated differential decreases in tolerant macrophage membrane G proteins. Gi3α was reduced the most to 48 ± 8% of controls (n = 3), and this reduction was significant compared to those of other G proteins. Gi1,2α and Gβ were reduced to 73 ± 5% (n = 3) and 65 ± 4% (n = 3) of control values, respectively. Gs α) (L) and Gs α(H) were reduced to 61 ± 5% (n = 3) and 68 ± 3% (n = 3) of control, respectively. These results demonstrate that endotoxin tolerant macrophages exhibit decreased membrane GTP binding capacity and differential reductions in the content of specific G proteins. The cellular mechanisms leading to such alterations in G proteins and their functional significance in the acquisition of endotoxin tolerance merit further investigation

Особливості хірургічного лікування вогнепальних поранень живота

Особливості хірургічного лікування вогнепальних поранень живот

Differential regulation of lipopolysaccharide and Gram-positive bacteria induced cytokine and chemokine production in splenocytes by Gαi proteins

AbstractHeterotrimeric Gi proteins play a role in lipopolysaccharide (LPS) and Staphylococcus aureus (SA) activated signaling leading to inflammatory mediator production. We hypothesized that genetic deletion of Gi proteins would alter cytokine and chemokine production induced by LPS and SA. LPS- and heat killed SA-induced cytokine and chemokine production in splenocytes from wild type (WT), Gαi2 (−/−) or Gαi1/3 (−/−) mice were investigated. LPS- or SA-induced production of TNFα, IL-6, IFNγ, IL-12, IL-17, GM-CSF, MIP-1α, MCP-1, MIG and IP-10 were significantly increased (1.2 to 33 fold, p<0.05) in splenocytes harvested from Gαi2(−/−) mice compared with WT mice. The effect of Gαi protein depletion was remarkably isoform specific. In splenocytes from Gαi1/3 (−/−) mice relative to WT mice, SA-induced IL-6, IFNγ, GM-CSF, and IP-10 levels were decreased (59% to 86%, p<0.05), whereas other LPS- or SA-stimulated cytokines and chemokines were not different relative to WT mice. LPS- and SA-induced production of KC were unchanged in both groups of the genetic deficient mice. Splenocytes from both Gαi2 (−/−) and Gαi1/3 (−/−) mice did not exhibit changes in TLR2 and TLR4 expression. Also analysis of splenic cellular composition by flow cytometry demonstrated an increase in splenic macrophages and reduced CD4 T cells in both Gαi2 (−/−) and Gαi1/3 (−/−) mice relative to WT mice. The disparate response of splenocytes from the Gαi2 (−/−) relative to Gαi1/3 (−/−) mice therefore cannot be attributed to major differences in spleen cellular composition. These data demonstrate that Gi2 and Gi1/3 proteins are both involved and differentially regulate splenocyte inflammatory cytokine and chemokine production in a highly Gi isoform specific manner in response to LPS and Gram-positive microbial stimuli