[Renin Angiotensin Axis, Angiotensin Converting Enzyme 2 and Coronavirus]. / Eje Renina Angiotensina, Enzima Convertidora de Angiotensina 2 y Coronavirus.

The renin-angiotensin-aldosterone system (RAAS) is the main plasma volume regulator, which maintains cardiovascular and hydrosaline homeostasis. In the classical pathway, the angiotensin converting enzyme (ACE) generates Angiotensin II (AngII), which is powerfully inflammatory and vasoconstrictive. This classical pathway is also regulated by ACE2, which converts AngI to Ang 1-9, and degrades AngII to Ang 1-7, whose vasodilatory and anti-inflammatory functions balance out the effects of AngII. ACE2 has been associated with the pathogenesis of respiratory infections such as RSV and severe acute respiratory syndrome coronavirus (SARS-CoV and SARS-CoV-2). Recent studies have shown that ACE2 corresponds to the main SARS-CoV-2 receptor, which together with other receptors such as the TMPRSS2, allows the virus to attach, fuse, and enter the host cell. These studies have shown that in animals infected with coronavirus there is a drop in tissue concentration of ACE2 and Ang 1-7, leading to overexpression of AngII and its vasoconstrictive and inflammatory effects. Experiments with recombinant ACE2 have shown a protective effect against overexpression of RAAS in coronavirus-infected animals, which is similar to that demonstrated with the use of AngII receptor blockers (AT1). Evidence on the protective role of ACE2 seems to support the recommendations re garding not discontinuing these drugs in COVID-19 infection. In this article, we present the current knowledge about the role of RAAS in coronavirus infection, based on physiopathological concepts, molecular bases, and experimental and clinical evidence.

Eje Renina Angiotensina, Enzima Convertidora de Angiotensina 2 y Coronavirus / Renin Angiotensin Axis, Angiotensin Converting Enzyme 2 and Coronavirus

Resumen: El sistema renina angiotensina aldosterona (SRAA) es el principal regulador del volumen plasmático, manteniendo la homeostasis cardiovascular e hidrosalina. En la vía clásica, la enzima convertidora de angiotensina (ECA) genera Angiotensina II (AngII), de potente efecto inflamatorio y vasoconstrictor. Esta vía clásica es a su vez regulada por la ECA2, que convierte AngII a Ang 1-7, cuyas acciones vaso dilatadoras y antiinflamatorias dan balance a los efectos de AngII. La ECA2 se ha relacionado con la patogenia de infecciones respiratorias como el virus respiratorio sincicial y el síndrome respiratorio agudo grave por coronavirus (SARS-CoV y SARS-CoV-2). Estudios recientes han demostrado que la ECA2 corresponde al principal receptor del SARS-CoV-2, que en conjunto con otros receptores como la serin proteasa TMPRSS2, permiten la fijación, fusión y entrada del virus a la célula huésped. En animales infectados por SARS-CoV se produce una caída de la concentración tisular de ECA2 y Ang 1-7, con la consiguiente sobreexpresión de AngII, y sus efectos vasoconstrictores e inflamatorios. Experimentos con ECA2 recombinante han mostrado un efecto protector frente a la sobreexpresión del SRAA en animales infectados por SARS-CoV, efecto similar al demostrado con el uso de bloquea- dores del receptor de AngII, AT1. La evidencia sobre el rol protector de ECA2 parece respaldar las recomendaciones respecto a no suspender estos medicamentos en la infección SARS-CoV-2. En este artículo presentamos el conocimiento actual sobre el rol del SRAA en la infección por SARS-CoV, a partir de conceptos fisiopatológicos, bases moleculares, y evidencia experimental y clínica.

Abstract: The renin-angiotensin-aldosterone system (RAAS) is the main plasma volume regulator, which maintains cardiovascular and hydrosaline homeostasis. In the classical pathway, the angiotensin converting enzyme (ACE) generates Angiotensin II (AngII), which is powerfully inflammatory and vasoconstrictive. This classical pathway is also regulated by ACE2, which converts AngI to Ang 1-9, and degrades AngII to Ang 1-7, whose vasodilatory and anti-inflammatory functions balance out the effects of AngII. ACE2 has been associated with the pathogenesis of respiratory infections such as RSV and severe acute respiratory syndrome coronavirus (SARS-CoV and SARS-CoV-2). Recent studies have shown that ACE2 corresponds to the main SARS-CoV-2 receptor, which together with other receptors such as the TMPRSS2, allows the virus to attach, fuse, and enter the host cell. These studies have shown that in animals infected with coronavirus there is a drop in tissue concentration of ACE2 and Ang 1-7, leading to overexpression of AngII and its vasoconstrictive and inflammatory effects. Experiments with recombinant ACE2 have shown a protective effect against overexpression of RAAS in coronavirus-infected animals, which is similar to that demonstrated with the use of AnglI receptor blockers (AT1). Evidence on the protective role of ACE2 seems to support the recommendations re garding not discontinuing these drugs in COVID-19 infection. In this article, we present the current knowledge about the role of RAAS in coronavirus infection, based on physiopathological concepts, molecular bases, and experimental and clinical evidence.

Scandinavian clinical practice guideline on fluid and drug therapy in adults with acute respiratory distress syndrome.

BACKGROUND: The objective of the Scandinavian Society of Anaesthesiology and Intensive Care Medicine (SSAI) task force on fluid and drug therapy in adults with acute respiratory distress syndrome (ARDS) was to provide clinically relevant, evidence-based treatment recommendations according to standards for trustworthy guidelines. METHODS: The guideline was developed according to standards for trustworthy guidelines, including a systematic review of the literature and use of the GRADE methodology for assessment of the quality of evidence and for moving from evidence to recommendations. RESULTS: A total of seven ARDS interventions were assessed. We suggest fluid restriction in patients with ARDS (weak recommendation, moderate quality evidence). Also, we suggest early use of neuromuscular blocking agents (NMBAs) in patients with severe ARDS (weak recommendation, moderate quality evidence). We recommend against the routine use of other drugs, including corticosteroids, beta2 agonists, statins, and inhaled nitric oxide (iNO) or prostanoids in adults with ARDS (strong recommendations: low- to high-quality evidence). These recommendations do not preclude the use of any drug or combination of drugs targeting underlying or co-existing disorders. CONCLUSION: This guideline emphasizes the paucity of evidence of benefit - and potential for harm - of common interventions in adults with ARDS and highlights the need for prudence when considering use of non-licensed interventions in this patient population.

Scandinavian clinical practice guideline on mechanical ventilation in adults with the acute respiratory distress syndrome

BACKGROUND: The objective of the Scandinavian Society of Anaesthesiology and Intensive Care Medicine (SSAI) task force on mechanical ventilation in adults with the acute respiratory distress syndrome (ARDS) is to formulate treatment recommendations based on available evidence from systematic reviews and randomised trials. METHODS: This guideline was developed according to standards for trustworthy guidelines through a systematic review of the literature and the use of the Grading of Recommendations Assessment, Development and Evaluation system for assessment of the quality of evidence and for moving from evidence to recommendations in a systematic and transparent process. RESULTS: We found evidence of moderately high quality to support a strong recommendation for pressure limitation and small tidal volumes in patients with ARDS. Also, we suggest positive end-expiratory pressure (PEEP) > 5 cm H2O in moderate to severe ARDS and prone ventilation 16/24 h for the first week in moderate to severe ARDS (weak recommendation, low quality evidence). Volume controlled ventilation or pressure control may be equally beneficial or harmful and partial modes of ventilatory support may be used if clinically feasible (weak recommendation, very low quality evidence). We suggest utilising recruitment manoeuvres as a rescue measure in catastrophic hypoxaemia only (weak recommendation, low quality evidence). Based on high-quality evidence, we strongly recommend not to use high-frequency oscillatory ventilation. We could find no relevant data from randomised trials to guide decisions on choice of FiO2 or utilisation of non-invasive ventilation. CONCLUSION: We strongly recommend pressure- and volume limitation and suggest using higher PEEP and prone ventilation in patients with severe respiratory failure.(AU)

Scandinavian clinical practice guideline on mechanical ventilation in adults with the acute respiratory distress syndrome.

BACKGROUND: The objective of the Scandinavian Society of Anaesthesiology and Intensive Care Medicine (SSAI) task force on mechanical ventilation in adults with the acute respiratory distress syndrome (ARDS) is to formulate treatment recommendations based on available evidence from systematic reviews and randomised trials. METHODS: This guideline was developed according to standards for trustworthy guidelines through a systematic review of the literature and the use of the Grading of Recommendations Assessment, Development and Evaluation system for assessment of the quality of evidence and for moving from evidence to recommendations in a systematic and transparent process. RESULTS: We found evidence of moderately high quality to support a strong recommendation for pressure limitation and small tidal volumes in patients with ARDS. Also, we suggest positive end-expiratory pressure (PEEP) > 5 cm H2O in moderate to severe ARDS and prone ventilation 16/24 h for the first week in moderate to severe ARDS (weak recommendation, low quality evidence). Volume controlled ventilation or pressure control may be equally beneficial or harmful and partial modes of ventilatory support may be used if clinically feasible (weak recommendation, very low quality evidence). We suggest utilising recruitment manoeuvres as a rescue measure in catastrophic hypoxaemia only (weak recommendation, low quality evidence). Based on high-quality evidence, we strongly recommend not to use high-frequency oscillatory ventilation. We could find no relevant data from randomised trials to guide decisions on choice of FiO2 or utilisation of non-invasive ventilation. CONCLUSION: We strongly recommend pressure- and volume limitation and suggest using higher PEEP and prone ventilation in patients with severe respiratory failure.

Modelling of hypoxaemia after gynaecological laparotomy.

BACKGROUND: Late postoperative arterial hypoxaemia is common after major surgery, and may contribute to cardiovascular, cerebral or wound complications. This study investigates the time course of hypoxaemia following gynaecological laparotomy, and estimates parameters of mathematical models of pulmonary gas exchange to describe hypoxaemia. METHODS: Twelve patients were studied on four occasions; preoperatively, 2, 8 and 48 h after surgery. On each occasion inspired oxygen fraction (FIO2) was varied, changing end-expired oxygen fraction (FEO2) to achieve arterial oxygen saturations (SaO2) ranging from 90% to 100%. Measurements of ventilation and blood gases were taken. Oxygenation was characterized plotting FEO2 against SaO2. The shape and position of the FEO2/SaO2 curve was described using two mathematical models including parameters describing gas exchange: either shunt and resistance to oxygen diffusion (Rdiff); or shunt and asymmetry of ventilation-perfusion (fA2). RESULTS: Two hours after surgery SaO2 was reduced from 97.5%+/-1.2% (mean+/-SD) to 93.8%+/-2.7% (mean+/-SD) (P<0.001). Values of shunt, Rdiff and fA2 were significantly changed at 2 and 8 h postoperatively. Forty-eight hours postoperatively Rdiff and fA2 were still significantly changed. CONCLUSION: Oxygenation in 12 patients preoperatively, 2, 8 and 48 h after gynaecological laparotomy is described. Two patients were hypoxaemic (SaO2 <92%) 48 h postoperatively. When two different models of oxygen transport are fitted to patient data, high values of Rdiff or low values of fA2 describe the right shift in the FEO2/SaO2 curve seen in patients with oxygenation problems. These models fit patient data identically, and may be useful in quantifying postoperative hypoxaemia.

Lrp binds to two regions in the dadAX promoter region of Escherichia coli to repress and activate transcription directly.

The dadAX operon is expressed by multiple promoters that are repressed by leucine-responsive regulatory protein (Lrp) and activated by cyclic AMP-CRP. In previous work, we found that alanine or leucine acted as inducers to antagonize Lrp repression of the three major promoters directly. Here, we identify 11 Lrp binding sites located within 350 bp of dad DNA. A mutational analysis, coupled with in vivo and in vitro transcription experiments, indicated that Lrp sites that overlap the dad promoters were involved in repression. In contrast, sites upstream of the promoters did not appear to be necessary for repression, but were required for activation by Lrp plus alanine or leucine of one of the major dad promoters, P2. This activation by alanine or leucine was not simply relief of repression, as P2 transcription from a constitutive template was increased fivefold compared with the basal level of transcription found in the absence of Lrp and the co-activator cyclic AMP-CRP. Alanine or leucine decreased the affinity of Lrp to repressor sites, while having little or no effect on the binding of Lrp to activator sites. This differential effect of alanine and leucine on Lrp binding helps to explain how these modifiers influence both repression and activation of the dad operon.

Delivering health and mental health care services to children in family foster care after welfare and health care reform.

As the 20th century draws to a close, fundamental changes in the organization, financing, and delivery of health care and welfare services, principally directed at poor families, are likely to result in an increased number of children entering out-of-home care. These children typically have significant physical, mental health, and developmental problems. Whether the quality of health care services they receive will improve as a result of health care reform efforts and new approaches to service delivery remains to be seen. This article addresses some of the major changes wrought by welfare and health care reform and describes the essential features of a health care system that can meet the special needs of children in care.

The case against preadoption genetic testing.

Adoption professionals and prospective adoptive families have become increasingly interested in obtaining genetic information on children prior to adoption. Predictive genetic testing of apparently healthy children has been urged as a way of generating information about children's future health and assisting families in deciding whether to adopt. Such genetic testing of children, however, raises a host of ethical issues with important implications for adoption policy and practice. The medical and psychosocial benefits and risks of predictive testing provide the context for analyzing the ethics of such testing. Ethical considerations strongly counsel against predictive genetic testing solely for purposes of evaluating a child for adoption.

In vitro and in vivo characterization of three major dadAX promoters in Escherichia coli that are regulated by cyclic AMP-CRP and Lrp.

We have shown previously that the dadAX operon of Escherichia coli expresses multiple transcripts, which are repressed by leucine-responsive regulatory protein (Lrp). Here we used site-directed mutagenesis and in vitro and in vivo transcription assays to show that each of the three major dad transcripts requires a specific promoter. These promoters, P1-P3, overlap and are positively regulated in vivo by cyclic AMP-CRP. DNase I footprinting experiments localized two CRP binding sites in this region: CRP1, which is positioned upstream of P1-P3, and CRP2, which is located within the promoters. Site-directed mutagenesis of each site provided evidence that CRP1 is necessary for the effects of cyclic AMP-CRP on dad expression in vivo and in vitro, and that CRP2 probably plays little or no role in this process.

Transcriptional activation by FNR and CRP: reciprocity of binding-site recognition.

Anaerobic expression of the focA pfl operon is dependent on the transcription factors ArcA and FNR and transcription is directed by multiple, anaerobically regulated promoters. A FNR-binding site is centred at -41.5 bp relative to the P6 promoter, inactivation of which severely impairs anaerobic expression of the complete operon. Mutations were introduced into this binding site to create a consensus recognition site for the cAMP-receptor protein, CRP (CC-site), and one that was recognised by both CRP and FNR (CF-site). Transcription directed by these mutant binding sites in vivo in different promoter constructs was analysed by primer extension and by constructing lacZ operon fusions. With a derivative including only the P6 promoter and the CF-binding site, transcription was shown to be independent of oxygen and was activated by CRP or FNR. In agreement with previous findings, FNR only activated transcription anaerobically. In a construct including the CC-binding site transcription was strong. CRP dependent and initiated at the identical site to the wild-type promoter. Transcription activation from the CC-site was exquisitely sensitive to low cAMP concentration. Surprisingly, in a crp mutant, anaerobically inducible, FNR-dependent transcription directed by the CC-site was detected, indicating that FNR can recognise a consensus CRP-binding site in vivo. A strain unable to synthesise CRP or FNR exhibited no transcription from the P6 promoter. Essentially the same results were observed in a series of constructs that also included the promoter P7 and its regulatory sequences. Evidence is also presented which demonstrates that CRP activates transcription from the natural FNR-binding site of the P6 promoter. In vitro DNA-binding studies showed that CRP specifically interacted with the FNR-binding site, protecting exactly the same sequence as that protected by the FNR protein. Interaction of CRP with the natural FNR-binding site was reduced greater than 50-fold compared to its interaction with the mutant CC-binding site. Although we could not demonstrate that FNR interacted with the CC-binding site in vitro, it did bind to the CF-site giving the same protection as observed with the wild-type FNR-binding site. FNR also activated transcription from the CF-site in vitro, giving further support to the idea that a single functional DNA half-site is sufficient to direct binding and transcription activation by a dimeric transcription factor.

Lrp is a direct repressor of the dad operon in Escherichia coli.

Expression of the degradative D-amino acid dehydrogenase (dad) operon is known to be increased when Escherichia coli is grown in the presence of D- or L-alanine. Alanine is thought to act as an inducer to block the action of a postulated repressor. This operon is also believed to be regulated by catabolite repression. We have used in vivo and in vitro experiments that show that the dad repressor is the leucine-responsive regulatory protein (Lrp). dad expression in a dad-lacZ operon fusion strain was increased four- to sevenfold when cells were grown in minimal medium containing alanine or leucine. A strain lacking Lrp had high-level constitutive dad expression. Gel retardation and footprinting studies revealed that Lrp binds in vitro to multiple sites over a large area in the dad promoter region. This binding was reduced by alanine or leucine. In vitro transcription assays, using a plasmid template and primer extension analysis, identified three major dad transcripts (Tr1, Tr2, and Tr3). The formation of these transcripts was differentially regulated by cyclic AMP-cyclic AMP receptor protein complex, and each was strongly repressed by Lrp. Alanine or leucine completely (for Tr1 and Tr2) or partially (for Tr3) reversed Lrp inhibition. Site-directed mutagenesis of an Lrp binding site strongly reduced Lrp binding and prevented Lrp repression of dad transcription in vivo and in vitro. Taken together, these results strongly suggest that Lrp and alanine or leucine act directly to repress and induce, respectively, transcription of the dad operon.

Integration host factor amplifies the induction of the aceBAK operon of Escherichia coli by relieving IclR repression.

A binding site for integration host factor (IHF) was identified upstream of the aceBAK promoter. Under inducing conditions, IHF activates aceB::lacZ expression by opposing IclR repression. In contrast, IHF has little effect on aceB::lacZ expression under repressing conditions. The ability of IHF to relieve repression under inducing but not repressing conditions allows this protein to amplify the induction of aceBAK.

Effect of a radiant heater on post-operative hypothermia: comparison with a reflective blanket.

Thirty patients with post-operative hypothermia following major surgery (thoracic, abdominal, orthopaedic) were allocated randomly to either active warming with a radiant heater (500 W) or passive rewarming with a reflective blanket. Rectal temperature, mean skin temperature (at four measuring sites), continuous haemoglobin saturation and shivering were measured for 2 h post-operatively. Although post-operative heat supply with a radiant heater resulted in faster rewarming, there were no differences between the two groups with respect to haemoglobin saturation and shivering.

Endothelin-1 and cerebral blood flow: influence of hypoxia, hypercapnia and indomethacin on circulating endothelin levels in healthy volunteers.

We investigated the effect of moderate (FiO2 13%) and light hypoxia (FiO2 17%) and hypercapnia (CO2 2-4%) with or without indomethacin on circulating levels of endothelin/endothelins (ET) and cerebral blood flow (CBF) in healthy volunteers. In protocol A, 23 subjects were exposed to moderate hypoxia. In protocol B, 29 subjects were randomized to one of four groups: (1) placebo, (2) indomethacin, (3) indomethacin+light hypoxia and (4) indomethacin+hypercapnia. Indomethacin was given as an intravenous bolus dose of 0.4mgkg-1 body weight followed by continuous infusion of 0.4mgkg-1h-1 for 6h. Two different FiO2 were chosen, light hypoxia in protocol B was chosen due to application of a known cerebral vasoconstrictor with unknown effect on cerebral autoregulation. We found, that moderate hypoxia (protocol A) induced a significant increase in CBF from 59.0 to 73.0 ml 100 g-1 brain tissue min-1 (p < 0.00005) with an increase in circulating levels of ET from 1.7 to 1.9fmol ml-1 plasma. However, this difference did not reach statistical significance (p = 0.14). We found, that indomethacin given intravenously (protocol B groups 2-3-4) significantly elevated circulating levels of ET from 2.1 to 3.9fmol ml-1 plasma (p < 0.00005) and decreased CBF from 60.5 to 39.5 ml 100g-1 brain tissue min-1 (p < 0.00005) compared to baseline values. Exposure to light hypoxia/hypercapnia in the indomethacin group increased CBF to values not significantly different from baseline values. Although there was no statistical correlation between ET and CBF with and without indomethacin, our results suggest that ET may be involved in the cerebral vasoconstriction produced by indomethacin given intravenously.

micF antisense RNA has a major role in osmoregulation of OmpF in Escherichia coli.

micF RNA, produced from a multicopy plasmid, was originally shown to be a major factor in negative osmoregulation of the OmpF outer membrane protein in Escherichia coli. However, subsequent experiments with a micF deletion strain suggested that chromosomal micF RNA was not a key component in this process. We report here that micF RNA is essential for the reduction in OmpF levels in cells grown in media of low-to-intermediate levels of osmolarity. Under these conditions, the amount of OmpF was reduced up to 60% in the parent strain while OmpF levels were not altered in the micF deletion mutant. In medium of higher osmolarity, OmpF synthesis was strongly inhibited in both strains. RNA measurements showed that micF RNA levels rose rapidly in cells grown in low-to-intermediate levels of osmolarity concomitant with the reduction in OmpF protein, while ompF mRNA decreased strongly only during high-osmolarity conditions. Taken together, these results strongly suggest that the negative osmoregulation of OmpF at low-to-intermediate osmolarity levels requires micF RNA and that this is masked at higher osmolarity by the known strong inhibition of OmpF transcription by OmpR. Results consistent with this model were also obtained by using procaine, a compound reported to inhibit ompF expression by a mechanism very similar to that involved in osmoregulation.

Integration host factor is required for anaerobic pyruvate induction of pfl operon expression in Escherichia coli.

The expression of the pyruvate formate-lyase gene (pfl) is induced by anaerobic growth, and this is increased further by growth in pyruvate. Previous work has shown that anaerobic induction is strongly dependent on the activator FNR and partially dependent on a second transcription factor, ArcA, while pyruvate induction only required FNR. Anaerobic and pyruvate regulation both require the presence of a 5' nontranslated regulatory sequence which spans approximately 500 bp of DNA. A mobility shift assay was developed to identify proteins that bind to this regulatory region. Several binding activities were separated by heparin agarose chromatography, and one of these activities was characterized and shown to be integration host factor (IHF). Mobility shift and DNase I footprinting experiments defined a single IHF binding site in the pfl promoter-regulatory region. With pfl-lacZ fusions, it could be shown that introduction of a himD mutation abolished pyruvate-dependent induction of anaerobic expression in vivo. The same result was observed when the pfl IHF binding site was mutated. In addition, the partial anaerobic induction of expression found in an fnr strain was completely blocked in an fnr himD double mutant and in an fnr IHF binding site double mutant. Taken together, these data suggest that IHF is necessary for both pyruvate induction and the anaerobic induction mediated by ArcA.