Photoacoustic imaging of the spatial distribution of oxygen saturation in an ischemia-reperfusion model in humans

Photoacoustic imaging (PAI) is a novel hybrid imaging technique that combines the advantages of optical and ultrasound imaging to produce hyperspectral images of the tissue. The feasibility of measuring oxygen saturation (sO₂) with PAI has been demonstrated pre-clinically, but has limited use in humans under conditions of ischemia and reperfusion. As an important step towards making PAI clinically available, we present a study in which PAI was used to estimate the spatial distribution of sO₂ in vivo during and after occlusion of the finger of eight healthy volunteers. The results were compared with a commercial oxygen saturation monitor based on diffuse reflectance spectroscopy. We here describe the capability of PAI to provide spatially resolved picture of the evolution of sO₂ during ischemia following vascular occlusion of a finger, demonstrating the clinical viability of PAI as a non-invasive diagnostic tool for diseases indicated by impaired microvascularization.     

Impact of intensified chemotherapy in metastatic pancreatic ductal adenocarcinoma (PDAC) in clinical routine in Europe

Background

Pancreatic ductal adenocarcinoma (PDAC) is associated with poor prognosis. Gemcitabine is the standard chemotherapy for patients with metastatic pancreatic adenocarcinoma (MPA). Randomized clinical trials evaluating intensified chemotherapies including FOLFIRINOX and nab-paclitaxel plus gemcitabine (NAB+GEM) have shown improvement in survival. Here, we have evaluated the efficacy of intensified chemotherapy versus gemcitabine monotherapy in real-life settings across Europe.

The management of infections in children in general practice in Sweden: a repeated 1-week diagnosis-prescribing study in 5 counties in 2000 and 2002

A diagnosis-prescribing study was performed in 5 Swedish counties during 1 week in November in 2000 and repeated in 2002. The aim of the present study was to analyse data for children 0-15y of age who consulted a general practitioner with symptoms of an infection. During the 2 weeks studied, 4049 children were consulted. Respiratory tract infections (RTI) were the predominant diagnoses, above all among the youngest children, while the proportion of urinary tract infections and skin infections increased with increasing age. Between the y 2000 and 2002, the proportion of children allocated the diagnosis streptococcal tonsillitis and pneumonia decreased (p<0.01 and p<0.001, respectively) while the proportion of common cold increased (p<0.001). Antibiotic prescribing decreased from 55% to 48% (p<0.001) for respiratory infections between the years studied. The only significant changes in type of antibiotics prescribed were the increase of isoxazolylpenicillins (p<0.001) used for skin infection and the decrease of macrolides (p=0.001). A diagnostic test was used in more than half of the consultations. Of children allocated a RTI diagnosis, 36% were prescribed antibiotics when a C-reactive protein test was performed compared to 58% in those not tested. Further studies are needed in general practice to determine the optimal use of near-patient tests in children with RTI.     

Innate immune receptor NOD2 promotes vascular inflammation and formation of lipid‐rich necrotic cores in hypercholesterolemic mice

Atherosclerosis is an inflammatory disease associated with the activation of innate immune TLRs and nucleotide‐binding oligomerization domain‐containing protein (NOD)‐like receptor pathways. However, the function of most innate immune receptors in atherosclerosis remains unclear. Here, we show that NOD2 is a crucial innate immune receptor influencing vascular inflammation and atherosclerosis severity. 10‐week stimulation with muramyl dipeptide (MDP), the NOD2 cognate ligand, aggravated atherosclerosis, as indicated by the augmented lesion burden, increased vascular inflammation and enlarged lipid‐rich necrotic cores in Ldlr^?/? mice. Myeloid‐specific ablation of NOD2, but not its downstream kinase, receptor‐interacting serine/threonine‐protein kinase 2, restrained the expansion of the lipid‐rich necrotic core in Ldlr^?/? chimeric mice. In vitro stimulation of macrophages with MDP enhanced the uptake of oxidized low‐density lipoprotein and impaired cholesterol efflux in concordance with upregulation of scavenger receptor A1/2 and downregulation of ATP‐binding cassette transporter A1. Ex vivo stimulation of human carotid plaques with MDP led to increased activation of inflammatory signaling pathways p38 MAPK and NF‐κB‐mediated release of proinflammatory cytokines. Altogether, this study suggests that NOD2 contributes to the expansion of the lipid‐rich necrotic core and promotes vascular inflammation in atherosclerosis.     

Microbial-induced meprin β cleavage in MUC2 mucin and a functional CFTR channel are required to release anchored small intestinal mucus

The mucus that covers and protects the epithelium of the intestine is built around its major structural component, the gel-forming MUC2 mucin. The gel-forming mucins have traditionally been assumed to be secreted as nonattached. The colon has a two-layered mucus system where the inner mucus is attached to the epithelium, whereas the small intestine normally has a nonattached mucus. However, the mucus of the small intestine of meprin β-deficient mice was now found to be attached. Meprin β is an endogenous zinc-dependent metalloprotease now shown to cleave the N-terminal region of the MUC2 mucin at two specific sites. When recombinant meprin β was added to the attached mucus of meprin β-deficient mice, the mucus was detached from the epithelium. Similar to meprin β-deficient mice, germ-free mice have attached mucus as they did not shed the membrane-anchored meprin β into the luminal mucus. The ileal mucus of cystic fibrosis (CF) mice with a nonfunctional cystic fibrosis transmembrane conductance regulator (CFTR) channel was recently shown to be attached to the epithelium. Addition of recombinant meprin β to CF mucus did not release the mucus, but further addition of bicarbonate rendered the CF mucus normal, suggesting that MUC2 unfolding exposed the meprin β cleavage sites. Mucus is thus secreted attached to the goblet cells and requires an enzyme, meprin β in the small intestine, to be detached and released into the intestinal lumen. This process regulates mucus properties, can be triggered by bacterial contact, and is nonfunctional in CF due to poor mucin unfolding.The gastrointestinal (GI) tract is protected from self-digestion and microbiota by mucus (1). This mucus is differently organized throughout the GI tract: the stomach and colon have a two-layered mucus system with an inner mucus layer attached to the epithelium and formed by stratified mucin sheets (2). This layer is 50–200 µm thick and impenetrable for bacteria, is constantly renewed by secreted mucins from the goblet cells, and further toward the lumen proteolytically converted into a nonattached and less dense outer mucus layer. This outer layer is the habitat and nutritional source for the commensal bacteria (2). In contrast, the small intestine has only one single mucus type that is not attached to the epithelium (3).The main structural component of the intestinal mucus is the heavily glycosylated polymeric MUC2 mucin which is densely packed inside the goblet cell and after secretion and a 1,000-fold expansion forms flat, net-like structures stacked into stratified mucin sheets in the colon (4). The same MUC2 mucin is processed differently in the small intestine where it appeared less organized but still filled the space between villi (3). This mucus was easily aspirated and penetrable to beads the size of bacteria (3). Bacteria could penetrate, but still the space between the villi was kept free of bacteria in the small intestine due to effective intestinal peristalsis, fast mucus renewal, and a high concentration of antibacterial peptides and proteins (3, 5). In fact, the structure of the small intestinal mucus as a nonattached and less organized layer mimics the situation for the outer colon mucus layer that has been shown to be generated from the inner mucus layer by proteolytic processing of the MUC2 mucin (2).Meprins are zinc-dependent metalloendopeptidases and belong to the astacin family (6). They comprise two homologous enzymes, meprin α and meprin β, where meprin α is the secreted and meprin β the membrane-tethered variant. They can form heterodimers (meprin α and β), homodimers (meprin β), and large oligomers (meprin α), forming one of the largest secreted protease complexes known. Although both enzymes share a common domain structure, they exhibit distinct features in substrate recognition and cleavage specificities. The enzymes can hydrolyze a broad variety of substrates, such as growth factors, peptide hormones, or compounds of the extracellular matrix like procollagen III, fibronectin, and tenascin-C (7–9). The meprin β enzyme is highly expressed in the enterocytes of the small intestine and is thereby localized close to the mucin networks (10).Cystic fibrosis (CF) is a severe disease that affects most of the mucus-producing organs of the body (11). The disease is caused by a nonfunctional cystic fibrosis transmembrane conductance regulator (CFTR) channel that normally mediates passive transport of chloride and bicarbonate ions (12, 13). Although a majority of the clinical CF symptoms can be attributed to stagnant mucus, the more precise link between the lack of CFTR function and mucus properties has been difficult to understand. We recently showed that the small intestinal mucus of CF mice, in contrast to the WT, was attached to the epithelium and impossible to aspirate (14). Although CF mice have only minor lung problems, their intestinal phenotype is similar to the human disease characterized by meconium ileus and distal intestinal obstruction syndrome (DIOS). When the CF mucus was secreted into a solution of ∼100 mM bicarbonate, the mucus was released from its attachment. Mucins are densely packed in the goblet cell granulae due to Ca²⁺ ions and low pH, and the role of bicarbonate is to remove the Ca²⁺ ions and increase the pH to allow for the >1,000-fold mucin expansion (4). When already formed mucus was treated with bicarbonate, the mucus was normalized and possible to aspirate, although its increased mucin density remained largely unaltered (14). This suggested that mucin attachment and expansion might be different phenomena and made us analyze this further.We have now found that meprin β is able to cleave the MUC2 mucin N terminus and that this is involved in the detachment of the mucus of the small intestine in a process controlled by bacteria and a functional CFTR channel. We thus describe a fundamental constitutive mechanism which involves an endogenous protease acting on the mucus network to alter its attached properties.     

In vitro combinations of red blood cell,plasma and platelet components evaluated by thromboelastography

Background

Thromboelastography is increasingly used to evaluate coagulation in massively bleeding patients. The aim of this study was to investigate how different combinations of blood components affect in vitro whole blood clotting measured by thromboelastography.

Materials and methods

Packed red blood cells, plasma and platelets from fresh and old blood components were mixed in vitro, in proportions of 4:4:1, 5:5:2, 8:4:1 and 2:1:0, and analysed with thromboelastography. For the ratio 4:4:1 the experiment was done at both 37 °C and 32 °C.

Results

Thromboelastography curves were within normal reference values for the blood component proportions of 4:4:1 and 5:5:2. For 8:4:1, the angle and maximal amplitude were reduced below normal values, indicating low levels of fibrinogen and/or platelets. For the 2:1:0 proportion, all parameters were affected resulting in severely impaired in vitro clot formation. The reaction-time, reflecting the coagulation factor-dependent, initial clot formation, was slightly increased at a low temperature. Prolonged storage of the components did not affect the curve.

Discussion

With the introduction of guidelines on the management of massive bleeding it is important to have tools for the assessment of the new protocols. In vitro evaluation of mixtures of packed red blood cells, plasma and platelets by thromboelastography may be relevant in the prediction of in vivo clot formation and haemostasis.     

Calcium and pH-dependent packing and release of the gel-forming MUC2 mucin

MUC2, the major colonic mucin, forms large polymers by N-terminal trimerization and C-terminal dimerization. Although the assembly process for MUC2 is established, it is not known how MUC2 is packed in the regulated secretory granulae of the goblet cell. When the N-terminal VWD1-D2-D'D3 domains (MUC2-N) were expressed in a goblet-like cell line, the protein was stored together with full-length MUC2. By mimicking the pH and calcium conditions of the secretory pathway we analyzed purified MUC2-N by gel filtration, density gradient centrifugation, and transmission electron microscopy. At pH 7.4 the MUC2-N trimer eluted as a single peak by gel filtration. At pH 6.2 with Ca(2+) it formed large aggregates that did not enter the gel filtration column but were made visible after density gradient centrifugation. Electron microscopy studies revealed that the aggregates were composed of rings also observed in secretory granulae of colon tissue sections. The MUC2-N aggregates were dissolved by removing Ca(2+) and raising pH. After release from goblet cells, the unfolded full-length MUC2 formed stratified layers. These findings suggest a model for mucin packing in the granulae and the mechanism for mucin release, unfolding, and expansion.     

Early release of interalveolar synechiae under general anesthesia through fiberscopic nasal intubation

This article presents a treatment strategy for early release of interalveolar synechiae, aiming to facilitate early oral feeding and prevent temporomandibular joint ankylosis.The treatment results of 2 patients with van der Woude syndrome were retrospectively studied. Both patients underwent early surgical release of interalveolar synechiae under general anesthesia through fiberscopic nasal intubation. The 2 patients were treated at the ages of 6 and 14 days, respectively. The interincisival distances increased from 5 and 6 mm preoperatively to 11 and 10 mm immediately after surgery. This was increased further to 25 and 20 mm at long-term follow-up (6 and 24 months).In conclusion, synechiae between the upper and lower jaws can be safely treated at a very early age under general anesthesia with fiberscopic nasotracheal intubation. The purpose of early intervention in these cases is to facilitate oral feeding and prevent temporomandibular joint ankylosis.