Engineered interleukin-6-derived cytokines recruit artificial receptor complexes and disclose CNTF signaling via the OSMR.

Ciliary neurotrophic factor (CNTF) activates cells via the non-signaling α-receptor CNTF receptor (CNTFR) and the two signaling ß-receptors glycoprotein 130 (gp130) and leukemia inhibitory factor receptor (LIFR). The CNTF derivate, Axokine, was protective against obesity and insulin resistance, but clinical development was halted by the emergence of CNTF antibodies. The chimeric cytokine IC7 used the framework of interleukin (IL-)6 with the LIFR-binding site from CNTF to activate cells via IL-6R:gp130:LIFR complexes. Similar to CNTF/Axokine, IC7 protected mice from obesity and insulin resistance. Here, we developed CNTF-independent chimeras that specifically target the IL-6R:gp130:LIFR complex. In GIL-6 and GIO-6, we transferred the LIFR binding site from LIF or OSM to IL-6, respectively. While GIO-6 signals via gp130:IL-6R:LIFR and gp130:IL-6R:OSMR complexes, GIL-6 selectively activates the IL-6R:gp130:LIFR receptor complex. By re-evaluation of IC7 and CNTF, we discovered the Oncostatin M receptor (OSMR) as an alternative non-canonical high-affinity receptor leading to IL-6R:OSMR:gp130 and CNTFR:OSMR:gp130 receptor complexes, respectively. The discovery of OSMR as an alternative high-affinity receptor for IC7 and CNTF designates GIL-6 as the first truly selective IL-6R:gp130:LIFR cytokine, whereas GIO-6 is a CNTF-free alternative for IC7.

Synthetic receptor platform to identify loss-of-function single nucleotide variants and designed mutants in the death receptor Fas/CD95.

Synthetic biology has emerged as a useful technology for studying cytokine signal transduction. Recently, we described fully synthetic cytokine receptors to phenocopy trimeric receptors such as the death receptor Fas/CD95. Using a nanobody as an extracellular-binding domain for mCherry fused to the natural receptor's transmembrane and intracellular domain, trimeric mCherry ligands were able to induce cell death. Among the 17,889 single nucleotide variants in the SNP database for Fas, 337 represent missense mutations that functionally remained largely uncharacterized. Here, we developed a workflow for the Fas synthetic cytokine receptor system to functionally characterize missense SNPs within the transmembrane and intracellular domain of Fas. To validate our system, we selected five functionally assigned loss-of-function (LOF) polymorphisms and included 15 additional unassigned SNPs. Moreover, based on structural data, 15 gain-of-function or LOF candidate mutations were additionally selected. All 35 nucleotide variants were functionally investigated through cellular proliferation, apoptosis and caspases 3 and 7 cleavage assays. Collectively, our results showed that 30 variants resulted in partial or complete LOF, while five lead to a gain-of-function. In conclusion, we demonstrated that synthetic cytokine receptors are a suitable tool for functional SNPs/mutations characterization in a structured workflow.

Bispecific soluble cytokine receptor-nanobody fusions inhibit Interleukin (IL-)6 trans-signaling and IL-12/23 or tumor necrosis factor (TNF) signaling.

At least 0.5% of people in the Western world develop inflammatory bowel disease (IBD). While antibodies that block tumor necrosis factor (TNF) α and Interleukin (IL-)23 have been approved for the treatment of IBD, IL-6 antibodies failed in the phase II clinical trial due to non-tolerable side effects. However, two clinical phase II studies suggest that inhibiting IL-6/soluble IL-6R (sIL-6R)-induced trans-signaling via the cytokine receptor gp130 benefit IBD patients with fewer adverse events. Here we develop inhibitors targeting a combination of IL-6/sIL-6R and TNF or IL-12/IL-23 signaling, named cs130-TNFVHHFc and cs130-IL-12/23VHHFc. Surface plasmon resonance experiments showed that recombinant cs130-TNFVHHFc and cs130-IL-12/23VHHFc bind with high affinity to IL-6/sIL-6R complexes and human TNFα (hTNFα) or IL-12/IL-23, respectively. Immunoprecipitation experiments have verified the higher ordered complex formation of the inhibitors with IL-6/sIL-6R and IL-12. We demonstrated that cs130-TNFVHHFc and cs130-IL-12/23VHHFc block IL-6/sIL-6R trans-signaling-induced proliferation and STAT3 phosphorylation of Ba/F3-gp130 cells, as well as hTNFα- or IL-23-induced signaling, respectively. In conclusion, cs130-TNFVHHFc and cs130-IL-12/23VHHFc represent a class of dimeric and bispecific chimeric cytokine inhibitors that consist of a soluble cytokine receptor fused to anti-cytokine nanobodies.

Respiratory syncytial virus-approved mAb Palivizumab as ligand for anti-idiotype nanobody-based synthetic cytokine receptors.

Synthetic cytokine receptors can modulate cellular functions based on an artificial ligand to avoid off-target and/or unspecific effects. However, ligands that can modulate receptor activity so far have not been used clinically because of unknown toxicity and immunity against the ligands. Here, we developed a fully synthetic cytokine/cytokine receptor pair based on the antigen-binding domain of the respiratory syncytial virus-approved mAb Palivizumab as a synthetic cytokine and a set of anti-idiotype nanobodies (AIPVHH) as synthetic receptors. Importantly, Palivizumab is neither cross-reactive with human proteins nor immunogenic. For the synthetic receptors, AIPVHH were fused to the activating interleukin-6 cytokine receptor gp130 and the apoptosis-inducing receptor Fas. We found that the synthetic cytokine receptor AIPVHHgp130 was efficiently activated by dimeric Palivizumab single-chain variable fragments. In summary, we created an in vitro nonimmunogenic full-synthetic cytokine/cytokine receptor pair as a proof of concept for future in vivo therapeutic strategies utilizing nonphysiological targets during immunotherapy.

A Hybrid Soluble gp130/Spike-Nanobody Fusion Protein Simultaneously Blocks Interleukin-6 trans-Signaling and Cellular Infection with SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can induce mild to life-threatening symptoms. Especially individuals over 60 years of age or with underlying comorbidities, including heart or lung disease and diabetes, or immunocompromised patients are at a higher risk. Fatal multiorgan damage in coronavirus disease 2019 (COVID-19) patients can be attributed to an interleukin-6 (IL-6)-dominated cytokine storm. Consequently, IL-6 receptor (IL-6R) monoclonal antibody treatment for severe COVID-19 cases has been approved for therapy. High concentrations of soluble IL-6R (sIL-6R) were found in COVID-19 intensive care unit patients, suggesting the involvement of IL-6 trans-signaling in disease pathology. Here, in analogy to bispecific antibodies (bsAbs), we developed the first bispecific IL-6 trans-signaling inhibitor, c19s130Fc, which blocks viral infection and IL-6 trans-signaling. c19s130Fc is a designer protein of the IL-6 trans-signaling inhibitor cs130 fused to a single-domain nanobody directed against the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. c19s130Fc binds with high affinity to IL-6:sIL-6R complexes as well as the spike protein of SARS-CoV-2, as shown by surface plasmon resonance. Using cell-based assays, we demonstrate that c19s130Fc blocks IL-6 trans-signaling-induced proliferation and STAT3 phosphorylation in Ba/F3-gp130 cells as well as SARS-CoV-2 infection and STAT3 phosphorylation in Vero cells. Taken together, c19s130Fc represents a new class of bispecific inhibitors consisting of a soluble cytokine receptor fused to antiviral nanobodies and principally demonstrates the multifunctionalization of trans-signaling inhibitors. IMPORTANCE The availability of effective SARS-CoV-2 vaccines is a large step forward in managing the pandemic situation. In addition, therapeutic options, e.g., monoclonal antibodies to prevent viral cell entry and anti-inflammatory therapies, including glucocorticoid treatment, are currently developed or in clinical use to treat already infected patients. Here, we report a novel dual-specificity inhibitor to simultaneously target SARS-CoV-2 infection and virus-induced hyperinflammation. This was achieved by fusing an inhibitor of viral cell entry with a molecule blocking IL-6, a key mediator of SARS-CoV-2-induced hyperinflammation. Through this dual action, this molecule may have the potential to efficiently ameliorate symptoms of COVID-19 in infected individuals.

Tryptophan (W) at position 37 of murine IL-12/IL-23 p40 is mandatory for binding to IL-12Rß1 and subsequent signal transduction.

Interleukin (IL)-12 and IL-23 are composite cytokines consisting of p35/p40 and p19/p40, respectively, which signal via the common IL-12 receptor ß1 (IL-12Rß1) and the cytokine-specific receptors IL-12Rß2 and IL-23R. Previous data showed that the p40 component interacts with IL-12Rß1, whereas p19 and p35 subunits solely bind to IL-23R and IL-12Rß2, resulting in tetrameric signaling complexes. In the absence of p19 and p35, p40 forms homodimers and may induce signaling via IL-12Rß1 homodimers. The critical amino acids of p19 and p35 required for binding to IL-23R and IL-12Rß2 are known, and two regions of p40 critical for binding to IL-12Rß1 have recently been identified. In order to characterize the involvement of the N-terminal region of p40 in binding to IL-12Rß1, we generated deletion variants of the p40-p19 fusion cytokine. We found that an N-terminal deletion variant missing amino acids M23 to P39 failed to induce IL-23-dependent signaling and did not bind to IL-12Rß1, whereas binding to IL-23R was maintained. Amino acid replacements showed that p40W37K largely abolished IL-23-induced signal transduction and binding to IL-12Rß1, but not binding to IL-23R. Combining p40W37K with D36K and T38K mutations eliminated the biological activity of IL-23. Finally, homodimeric p40D36K/W37K/T38K did not interact with IL-12Rß1, indicating binding of homodimeric p40 to IL-12Rß1 is comparable to the interaction of IL-23/IL-12 and IL-12Rß1. In summary, we have defined D36, W37, and T38 as hotspot amino acids for the interaction of IL-12/IL-23 p40 with IL-12Rß1. Structural insights into cytokine-cytokine receptor binding are important to develop novel therapeutic strategies.

Immunoreceptor Engineering and Synthetic Cytokine Signaling for Therapeutics.

Cytokines control immune-related events and are critically involved in a plethora of physiological and pathophysiological processes including autoimmunity and cancer development. Accordingly, modulation of natural cytokine signaling by antibodies and small molecules has improved therapeutic regimens. Synthetic biology sets out to optimize immunotherapeutics, with chimeric antigen receptor (CAR) T cell immmunotherapy being the first example to combine synthetic biology with genetic engineering during therapy. Hence, synthetic cytokines and cytokine receptors, as well as constitutively active cytokine receptor variants, are emerging as tools to improve or modulate immunotherapeutic strategies. This review focuses on recent developments in the growing field of synthetic cytokine signaling, providing an outlook for developing applications that involve physiological targets of immunotherapy.

Deciphering site 3 interactions of interleukin 12 and interleukin 23 with their cognate murine and human receptors.

Interleukin (IL)-12 and IL-23 belong to the IL-12 type family and are composite cytokines, consisting of the common ß subunit p40 and the specific cytokine α subunit p35 and p19, respectively. IL-12 signals via the IL-12Rß1·IL-12Rß2 receptor complex, and IL-23 uses also IL-12Rß1 but engages IL-23R as second receptor. Importantly, binding of IL-12 and IL-23 to IL-12Rß1 is mediated by p40, and binding to IL-12Rß2 and IL-23R is mediated by p35 and p19, respectively. Previously, we have identified a W157A substitution at site 3 of murine IL-23p19 that abrogates binding to murine IL-23R. Here, we demonstrate that the analogous Y185R site 3 substitution in murine and Y189R site 3 substitution in human IL-12p35 abolishes binding to IL-12Rß2 in a cross-species manner. Although Trp157 is conserved between murine and human IL-23p19 (Trp156 in the human ortholog), the site 3 W156A substitution in hIL-23p19 did not affect signaling of cells expressing human IL-12Rß1 and IL-23R, suggesting that the interface of murine IL-23p19 required for binding to IL-23R is different from that in the human ortholog. Hence, we introduced additional hIL-23p19 substitutions within its binding interface to hIL-23R and found that the combined site 3 substitutions of W156A and L160E, which become buried at the complex interface, disrupt binding of hIL-23p19 to hIL-23R. In summary, we have identified substitutions in IL-12p35 and IL-23p19 that disrupt binding to their cognate receptors IL-12Rß2 and IL-23R in a murine/human cross-species manner.

Current status and relevance of single nucleotide polymorphisms in IL-6-/IL-12-type cytokine receptors.

Cytokines control immune related events and are critically involved in a plethora of patho-physiological processes including autoimmunity and cancer development. In rare cases, single nucleotide polymorphisms (SNPs) or single nucleotide variations (SNVs) in cytokine receptors eventually cause detrimental ligand-independent, constitutive activation of signal transduction. Most SNPs have, however, no or only marginal influences on gene expression, protein stability, localization and function and thereby only slightly affecting pathogenesis probability. The SNP database (dbSNP) is an archive for a broad collection of polymorphisms in which SNPs are categorized and marked with a locus accession number "reference SNP" (rs). Here, we engineered an algorithm to directly align dbSNP information to DNA and protein sequence information to clearly illustrate a genetic SNP landscape exemplified for all tall cytokine receptors of the IL-6/IL-12 family, including IL-23R, IL-12Rß1, IL-12Rß2, gp130, LIFR, OSMR and WSX-1. This information was complemented by a comprehensive literature summary and structural insights of relevant disease-causing SNPs in cytokine/cytokine receptor interfaces. In summary, we present a general strategy with potential to apply to other cytokine receptor networks.

IL-6 Trans-signaling Controls Liver Regeneration After Partial Hepatectomy.

Interleukin-6 (IL-6) is critically involved in liver regeneration after partial hepatectomy (PHX). Previous reports suggest that IL-6 trans-signaling through the soluble IL-6/IL-6R complex is involved in this process. However, the long-term contribution of IL-6 trans-signaling for liver regeneration after PHX is unknown. PHX-induced generation of the soluble IL-6R by ADAM (a disintegrin and metallo) proteases enables IL-6 trans-signaling, in which IL-6 forms an agonistic complex with the soluble IL-6 receptor (sIL-6R) to activate all cells expressing the signal-transducing receptor chain glycoprotein 130 (gp130). In contrast, without activation of ADAM proteases, IL-6 in complex with membrane-bound IL-6R and gp130 activates classic signaling. Here, we describe the generation of IL-6 trans-signaling mice, which exhibit boosted IL-6 trans-signaling and abrogated classic signaling by genetic conversion of all membrane-bound IL-6R into sIL-6R proteins phenocopying hyperactivation of ADAM-mediated shedding of IL-6R as single substrate. Importantly, although IL-6R deficient mice were strongly affected by PHX, survival and regeneration of IL-6 trans-signaling mice was indistinguishable from control mice, demonstrating that IL-6 trans-signaling fully compensates for disabled classic signaling in liver regeneration after PHX. Moreover, we monitored the long-term consequences of global IL-6 signaling inhibition versus IL-6 trans-signaling selective blockade after PHX by IL-6 monoclonal antibodies and soluble glycoprotein 130 as fragment crystallizable fusion, respectively. Both global IL-6 blockade and selective inhibition of IL-6 trans-signaling results in a strong decrease of overall survival after PHX, accompanied by decreased signal transducer and activator of transcription 3 phosphorylation and proliferation of hepatocytes. Mechanistically, IL-6 trans-signaling induces hepatocyte growth factor production by hepatic stellate cells. Conclusion: IL-6 trans-signaling, but not classic signaling, controls liver regeneration following PHX.

The balance of interleukin (IL)-6, IL-6·soluble IL-6 receptor (sIL-6R), and IL-6·sIL-6R·sgp130 complexes allows simultaneous classic and trans-signaling.

Interleukin (IL-)6 is the major pro-inflammatory cytokine within the IL-6 family. IL-6 signals via glycoprotein 130 (gp130) and the membrane-bound or soluble IL-6 receptor (IL-6R), referred to as classic or trans-signaling, respectively. Whereas inflammation triggers IL-6 expression, eventually rising to nanogram/ml serum levels, soluble IL-6R (sIL-6R) and soluble gp130 (sgp130) are constitutively present in the upper nanogram/ml range. Calculations based on intermolecular affinities have suggested that systemic IL-6 is immediately trapped in IL-6·sIL-6R and IL-6·sIL-6R·sgp130 complexes, indicating that sIL-6R and sgp130 constitute a buffer system that increases the serum half-life of IL-6 or restricts systemic IL-6 signaling. However, this scenario has not been experimentally validated. Here, we quantified IL-6·sIL-6R and IL-6·sIL-6R·sgp130 complexes over a wide concentration range. The amounts of IL-6 used in this study reflect concentrations found during active inflammatory events. Our results indicated that most IL-6 is free and not complexed with sIL-6R or sgp130, indicating that the level of endogenous sgp130 in the bloodstream is not sufficient to block IL-6 trans-signaling via sIL-6R. Importantly, addition of the single-domain antibody VHH6, which specifically stabilizes IL-6·sIL-6R complexes but did not bind to IL-6 or sIL-6R alone, drove free IL-6 into IL-6·sIL-6R complexes and boosted trans-signaling but not classic signaling, demonstrating that endogenous sIL-6R has at least the potential to form complexes with IL-6. Our findings indicate that even though high concentrations of sIL-6R and sgp130 are present in human serum, the relative ratio of free IL-6 to IL-6·sIL-6R allows for simultaneous classic and trans-signaling.

Titin-Based Cardiac Myocyte Stiffening Contributes to Early Adaptive Ventricular Remodeling After Myocardial Infarction.

RATIONALE: Myocardial infarction (MI) increases the wall stress in the viable myocardium and initiates early adaptive remodeling in the left ventricle to maintain cardiac output. Later remodeling processes include fibrotic reorganization that eventually leads to cardiac failure. Understanding the mechanisms that support cardiac function in the early phase post MI and identifying the processes that initiate transition to maladaptive remodeling are of major clinical interest. OBJECTIVE: To characterize MI-induced changes in titin-based cardiac myocyte stiffness and to elucidate the role of titin in ventricular remodeling of remote myocardium in the early phase after MI. METHODS AND RESULTS: Titin properties were analyzed in Langendorff-perfused mouse hearts after 20-minute ischemia/60-minute reperfusion (I/R), and mouse hearts that underwent ligature of the left anterior descending coronary artery for 3 or 10 days. Cardiac myocyte passive tension was significantly increased 1 hour after ischemia/reperfusion and 3 and 10 days after left anterior descending coronary artery ligature. The increased passive tension was caused by hypophosphorylation of the titin N2-B unique sequence and hyperphosphorylation of the PEVK (titin domain rich in proline, glutamate, valine, and lysine) region of titin. Blocking of interleukine-6 before left anterior descending coronary artery ligature restored titin-based myocyte tension after MI, suggesting that MI-induced titin stiffening is mediated by elevated levels of the cytokine interleukine-6. We further demonstrate that the early remodeling processes 3 days after MI involve accelerated titin turnover by the ubiquitin-proteasome system. CONCLUSIONS: We conclude that titin-based cardiac myocyte stiffening acutely after MI is partly mediated by interleukine-6 and is an important mechanism of remote myocardium to adapt to the increased mechanical demands after myocardial injury.

A modified CO2/O2 Guedel airway improves capnographic accuracy compared with a CO2/O2 nasal cannula: An infant manikin study.

BACKGROUND: Capnography via a CO2/O2 nasal cannula is commonly used for respiratory monitoring during sedation. However, signal disturbances are frequently encountered, especially in young children. OBJECTIVE: Sampling ports placed closer to the trachea have been shown to result in improved signal quality. In a manikin model of a 6-month-old infant we compared capnography from a modified Guedel airway with a CO2 port located at the tip with that from a CO2/O2 nasal cannula. DESIGN: A comparison study using an artificial model of a breathing 6-month-old infant. SETTING: Department of Paediatrics, Inselspital Bern, Switzerland, from March 2016 to June 2016. MATERIAL: Modified CO2/O2 Guedel airway. INTERVENTIONS: Capnography using a modified CO2/O2 Guedel airway or a CO2/O2 nasal cannula was performed for tidal volumes of 20 to 80 ml (in steps of 20 ml), respiratory rates of 20 to 60 min (in steps of 10 min) and with different O2 flows (0 to 2 l min, in steps of 0.5 l). MAIN OUTCOME MEASURES: Comparison of differences between tracheal and device CO2. Secondary outcomes included the effect of various respiratory settings and O2 flows on the CO2 difference. RESULTS: The tracheal to device CO2 difference was significantly smaller when using a modified CO2/O2 Guedel airway vs. a CO2/O2 nasal cannula: Mean ±â€ŠSD, 16.8 ±â€Š4.9 vs. 24.1 ±â€Š5.9 mmHg, P less than 0.0001. An O2 flow of 0.5 to 2 l min did not influence the tracheal to device CO2 difference with the modified CO2/O2 Guedel airway in contrast to the CO2/O2 nasal cannula where there were significant differences (P < 0.0001). The effect of various tidal volumes and respiratory rates proved to be similar in both devices. CONCLUSION: Capnography traces derived from a sample port at the tip of a modified CO2/O2 Guedel airway were more accurate than those obtained from a CO2/O2 nasal cannula. TRIAL REGISTRATION: Not applicable.

Impact of high concentrations of sevoflurane on laryngeal reflex responses.

BACKGROUND: Exaggerated defensive upper airway reflexes, particularly laryngospasm, may cause hypoxemic damage, especially in children. General clinical experience suggests that laryngeal reflex responses are more common under light levels of anesthesia, and previous clinical studies have shown an inverse correlation between laryngeal responsiveness and depth of hypnosis. However, this seems to be less obvious in children anesthetized with sevoflurane. The aim of this study was to assess the impact of high concentrations of sevoflurane on laryngeal and respiratory reflex responses in spontaneously breathing children. Accordingly, we tested the hypothesis that laryngeal and respiratory reflex responses were completely suppressed in spontaneously breathing children when anesthetized with sevoflurane 4.7% (=MACED95Intubation ) as compared with sevoflurane 2.5% (=1 MAC). METHODS: In this prospective observational study, we tested the hypothesis that the incidence of laryngospasm evoked by laryngeal stimulation is diminished under high concentrations of sevoflurane. Following Ethics approval, trial registration, and informed consent, 40 children (3-7 years) scheduled for elective surgery participated in the trial. All children received sevoflurane 2.5% (1 MAC) and 4.7% (ED95Intubation ) in random order with 5-min equilibration between the states. Under both conditions, distilled water was sprayed under bronchoscopic view onto the larynx. Potential laryngeal and respiratory reflex responses were assessed offline by a blinded reviewer. RESULTS: Laryngospasm (episodes lasting >10 s) occurred in 12/38 (32%) of the patients anesthetized with sevoflurane 2.5%, vs 7/38 (18%) in those anesthetized with sevoflurane 4.7% (difference: OR 3.5; 95% CI [0.72-16.84], P = 0.18). All other reflex responses (coughing, expiration reflexes, and spasmodic panting) were infrequent and were similar among the examined concentrations. CONCLUSION: Against our hypothesis, laryngospasm could still be observed in 18% of children under the higher concentration of sevoflurane (4.7%, ED95Intubation ).

Non-canonical interleukin 23 receptor complex assembly: p40 protein recruits interleukin 12 receptor ß1 via site II and induces p19/interleukin 23 receptor interaction via site III.

IL-23, composed of the cytokine subunit p19 and the soluble α receptor subunit p40, binds to a receptor complex consisting of the IL-23 receptor (IL-23R) and the IL-12 receptor ß1 (IL-12Rß1). Complex formation was hypothesized to follow the "site I-II-III" architectural paradigm, with site I of p19 being required for binding to p40, whereas sites II and III of p19 mediate binding to IL-12Rß1 and IL-23R, respectively. Here we show that the binding mode of p19 to p40 and of p19 to IL-23R follow the canonical site I and III paradigm but that interaction of IL-23 to IL-12Rß1 is independent of site II in p19. Instead, binding of IL-23 to the cytokine binding module of IL-12Rß1 is mediated by domains 1 and 2 of p40 via corresponding site II amino acids of IL-12Rß1. Moreover, domains 2 and 3 of p40 were sufficient for complex formation with p19 and to induce binding of p19 to IL-23R. The Fc-tagged fusion protein of p40_D2D3/p19 did, however, not act as a competitive IL-23 antagonist but, at higher concentrations, induced proliferation via IL-23R but independent of IL-12Rß1. On the basis of our experimental validation, we propose a non-canonical topology of the IL-23·IL-23R·IL-12Rß1 complex. Furthermore, our data help to explain why p40 is an antagonist of IL-23 and IL-12 signaling and show that site II of p19 is dispensable for IL-23 signaling.

[Anesthesia for ophthalmic surgery]. / Anästhesie bei ophthalmologischen Operationen - eine Standortbestimmung.

The majority of ophthalmic interventions can be done today under locoregional anesthesia using "Monitored Anesthesia Care" (MAC). General anesthesia techniques are mostly reserved for the pediatric segment and for patients with specific comorbidity and/or lengthy procedure. Cataract surgery in predominantly geriatric patients belongs to the field of the so-called "high volume-surgery": Given the low perioperative risk in this patient group, adapted and optimized processes are indicated. A focused premedication and informing these patients ensures good perioperative compliance. Preoperative tests are be conducted in this patient population only as a function of relevant comorbidity. Premedication usually takes place as a classical anesthesia consultation, but new methods such as an internet-based premedication for healthy patients offers a new option. The intraoperative anesthesia method depends on the needs of the surgeon and the expectations and possibilities of cooperation of the patient.

Alternative intronic polyadenylation generates the interleukin-6 trans-signaling inhibitor sgp130-E10.

Interleukin (IL)-6 signals via a receptor complex composed of the signal-transducing ß-receptor gp130 and the non-signaling membrane-bound or soluble IL-6 receptor α (IL-6R, sIL-6R), which is referred to as classic and trans-signaling, respectively. IL-6 trans-signaling is functionally associated with the development of chronic inflammatory diseases and cancer. Soluble gp130 (sgp130) variants are natural inhibitors of trans-signaling. Differential splicing yields sgp130 isoforms. Here, we describe that alternative intronic polyadenylation in intron 10 of the gp130 transcript results in a novel mRNA coding for an sgp130 protein isoform (sgp130-E10) of 70-80 kDa. The sgp130-E10 protein was expressed in vivo in human peripheral blood mononuclear cells. To assess the biological activity of sgp130-E10, we expressed this variant as Fc-tagged fusion protein (sgp130-E10Fc). Recombinant sgp130-E10Fc binds to a complex of IL-6 and sIL-6R, but not to IL-6 alone, and specifically inhibits IL-6 trans-signaling. Thus, it might play an important role in the regulation of trans-signaling in vivo.

Assessment of three placement techniques for individualized positioning of the tip of the tracheal tube in children under the age of 4 years.

BACKGROUND: Accurate positioning of the tip of the tracheal tube (tube tip) is challenging in young children. Prevalent clinical methods include placement of intubation depth marks, palpation of the tube cuff in the suprasternal notch, or deliberate mainstem intubation with subsequent withdrawal. To compare the predictability of tube tip positions, variability of the resulting positions in relation to the carina was determined applying the three techniques in each patient. METHODS: In 68 healthy children aged ≤4 years, intubation was performed with an age-adapted, high-volume low-pressure cuffed tube adjusting the imprinted depth mark to the level of the vocal cords. The tube tip-to-carina distance was measured endoscopically. Thereafter, placements using (I) cuff palpation in the suprasternal notch and (II) auscultation to determine change in breath sounds during withdrawal after bronchial mainstem intubation were completed in random order. RESULTS: Tube tip position above the carina was higher when using depth marks (mean = 36.8 mm) compared with cuff palpation in the suprasternal notch (mean = 19.0 mm). Variability, expressed as sd, was lowest with the mainstem intubation technique (5.2 mm) followed by the cuff palpation (7.4 mm) and the depth mark technique (11.2 mm) (P < 0.005). CONCLUSION: Auscultation after deliberate mainstem intubation and cuff palpation resulted in a tube tip position above the carina that was shorter and more predictable than placement of the tube using depth markings.

Rho-kinase: regulation, (dys)function, and inhibition.

In a variety of normal and pathological cell types, Rho-kinases I and II (ROCKI/II) play a pivotal role in the organization of the nonmuscle and smooth muscle cytoskeleton and adhesion plaques as well as in the regulation of transcription factors. Thus, ROCKI/II activity regulates cellular contraction, motility, morphology, polarity, cell division, and gene expression. Emerging evidence suggests that dysregulation of the Rho-ROCK pathways at different stages is linked to cardiovascular, metabolic, and neurodegenerative diseases as well as cancer. This review focuses on the current status of understanding the multiple functions of Rho-ROCK signaling pathways and various modes of regulation of Rho-ROCK activity, thereby orchestrating a concerted functional response.

Targeting IL-6 trans-signalling: past, present and future prospects.

Interleukin-6 (IL-6) is a key immunomodulatory cytokine that affects the pathogenesis of diverse diseases, including autoimmune diseases, chronic inflammatory conditions and cancer. Classical IL-6 signalling involves the binding of IL-6 to the membrane-bound IL-6 receptor α-subunit (hereafter termed 'mIL-6R') and glycoprotein 130 (gp130) signal-transducing subunit. By contrast, in IL-6 trans-signalling, complexes of IL-6 and the soluble form of IL-6 receptor (sIL-6R) signal via membrane-bound gp130. A third mode of IL-6 signalling - known as cluster signalling - involves preformed complexes of membrane-bound IL-6-mIL-6R on one cell activating gp130 subunits on target cells. Antibodies and small molecules have been developed that block all three forms of IL-6 signalling, but in the past decade, IL-6 trans-signalling has emerged as the predominant pathway by which IL-6 promotes disease pathogenesis. The first selective inhibitor of IL-6 trans-signalling, sgp130, has shown therapeutic potential in various preclinical models of disease and olamkicept, a sgp130Fc variant, had promising results in phase II clinical studies for inflammatory bowel disease. Technological developments have already led to next-generation sgp130 variants with increased affinity and selectivity towards IL-6 trans-signalling, along with indirect strategies to block IL-6 trans-signalling. Here, we summarize our current understanding of the biological outcomes of IL-6-mediated signalling and the potential for targeting this pathway in the clinic.